genejet pcr purification kit  (Thermo Fisher)


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    Name:
    GeneJET PCR Purification Kit
    Description:
    Thermo Scientific GeneJET PCR Purification Kit utilizes a proprietary silica-based membrane technology in the form of a convenient spin column, eliminating the need for tedious resin manipulations or toxic phenol-chloroform extractions. It effectively removes primers, dNTPs, unincorporated labeled nucleotides, enzymes, and salts from PCR and other reaction mixtures.The kit can be used for purification of DNA fragments from 25 bp to 20 kb with recovery rates up to 100%. Each GeneJET purification column has a total binding capacity of up to 25 µg of DNA, and the entire procedure takes 5 minutes. The purified DNA can be used in common downstream applications, such as sequencing, restriction digestion, labeling, ligation, cloning, in vitro transcription, blotting or in situ hybridization.Highlights• Highly efficient: 90 to 100% recoveries in the range of 100 bp to 10 kb• Fast: procedure takes only 5 minutes• Convenient: spin columns are capped and assembled with collection tubesApplications• Fast and efficient purification of DNA fragments ideal for use in all conventional molecular biology procedures including:• FastDigest or conventional restriction digestion• Automated fluorescent or radioactive sequencing• PCR• In vitro transcription• In situ hybridization• Labeling• Cloning
    Catalog Number:
    K0701
    Price:
    None
    Applications:
    DNA & RNA Purification & Analysis|DNA Extraction|PCR Product Clean-Up
    Size:
    50 preps
    Category:
    Kits and Assays, DNA⁄RNA Purification Kits & Reagents, PCR Clean-Up Kits
    Score:
    85
    Buy from Supplier


    Structured Review

    Thermo Fisher genejet pcr purification kit
    DNA purification reagents vary in their ability to recover low amounts of DNA from de-crosslinked chromatin. a Recovered DNA amount by different DNA purification reagents from de-crosslinked chromatin. De-crosslinked chromatin estimated to include 1 ng range DNA in ChIP elution buffer was purified following the manufacturer’s instructions. The data were generated from triplicate DNA samples derived from three independent preparations. Zy, ChIP DNA Clean Concentrator™ (Zymo Research); Pr, Wizard® SV Gel and <t>PCR</t> Clean-Up System (Promega); Th, <t>GeneJET</t> PCR Purification Kit (Thermo Fisher Scientific); In, PureLink® PCR Purification Kit (Invitrogen); Ne, Monarch® PCR DNA Cleanup Kit (New England Biolabs); Am, Chromatin IP DNA Purification Kit (Active Motif); Qp, QIAquick PCR Purification Kit (Qiagen); Qm, MinElute PCR Purification Kit (Qiagen); Ba, Agencourt AMPure XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); Br, RNAClean™ XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); PC, phenol/chloroform extraction. b Interference of PCR amplification by purified eluent of purification reagents. 9 μL eluent was mixed with 1 μL 166 bp of Drosophila probe DNA (0.0001 ng), and the resulting mixture was used as the template in 20 μl of real-time PCR reaction. The Ct value for Drosophila probe DNA from TE buffer was set as 100%. The experiment was repeated 3 times using de-crosslinked chromatin estimated to include 1 ng of DNA. c Size profiles of DNA purified by different reagents. The DNAs purified from de-crosslinked chromatin estimated to include 50 ng range DNA was analyzed by AATI Fragment Analyzer. DNA size (bp) is shown
    Thermo Scientific GeneJET PCR Purification Kit utilizes a proprietary silica-based membrane technology in the form of a convenient spin column, eliminating the need for tedious resin manipulations or toxic phenol-chloroform extractions. It effectively removes primers, dNTPs, unincorporated labeled nucleotides, enzymes, and salts from PCR and other reaction mixtures.The kit can be used for purification of DNA fragments from 25 bp to 20 kb with recovery rates up to 100%. Each GeneJET purification column has a total binding capacity of up to 25 µg of DNA, and the entire procedure takes 5 minutes. The purified DNA can be used in common downstream applications, such as sequencing, restriction digestion, labeling, ligation, cloning, in vitro transcription, blotting or in situ hybridization.Highlights• Highly efficient: 90 to 100% recoveries in the range of 100 bp to 10 kb• Fast: procedure takes only 5 minutes• Convenient: spin columns are capped and assembled with collection tubesApplications• Fast and efficient purification of DNA fragments ideal for use in all conventional molecular biology procedures including:• FastDigest or conventional restriction digestion• Automated fluorescent or radioactive sequencing• PCR• In vitro transcription• In situ hybridization• Labeling• Cloning
    https://www.bioz.com/result/genejet pcr purification kit/product/Thermo Fisher
    Average 99 stars, based on 453 article reviews
    Price from $9.99 to $1999.99
    genejet pcr purification kit - by Bioz Stars, 2019-12
    99/100 stars

    Images

    1) Product Images from "Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application"

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application

    Journal: BMC Genomics

    doi: 10.1186/s12864-017-4371-5

    DNA purification reagents vary in their ability to recover low amounts of DNA from de-crosslinked chromatin. a Recovered DNA amount by different DNA purification reagents from de-crosslinked chromatin. De-crosslinked chromatin estimated to include 1 ng range DNA in ChIP elution buffer was purified following the manufacturer’s instructions. The data were generated from triplicate DNA samples derived from three independent preparations. Zy, ChIP DNA Clean Concentrator™ (Zymo Research); Pr, Wizard® SV Gel and PCR Clean-Up System (Promega); Th, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, PureLink® PCR Purification Kit (Invitrogen); Ne, Monarch® PCR DNA Cleanup Kit (New England Biolabs); Am, Chromatin IP DNA Purification Kit (Active Motif); Qp, QIAquick PCR Purification Kit (Qiagen); Qm, MinElute PCR Purification Kit (Qiagen); Ba, Agencourt AMPure XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); Br, RNAClean™ XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); PC, phenol/chloroform extraction. b Interference of PCR amplification by purified eluent of purification reagents. 9 μL eluent was mixed with 1 μL 166 bp of Drosophila probe DNA (0.0001 ng), and the resulting mixture was used as the template in 20 μl of real-time PCR reaction. The Ct value for Drosophila probe DNA from TE buffer was set as 100%. The experiment was repeated 3 times using de-crosslinked chromatin estimated to include 1 ng of DNA. c Size profiles of DNA purified by different reagents. The DNAs purified from de-crosslinked chromatin estimated to include 50 ng range DNA was analyzed by AATI Fragment Analyzer. DNA size (bp) is shown
    Figure Legend Snippet: DNA purification reagents vary in their ability to recover low amounts of DNA from de-crosslinked chromatin. a Recovered DNA amount by different DNA purification reagents from de-crosslinked chromatin. De-crosslinked chromatin estimated to include 1 ng range DNA in ChIP elution buffer was purified following the manufacturer’s instructions. The data were generated from triplicate DNA samples derived from three independent preparations. Zy, ChIP DNA Clean Concentrator™ (Zymo Research); Pr, Wizard® SV Gel and PCR Clean-Up System (Promega); Th, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, PureLink® PCR Purification Kit (Invitrogen); Ne, Monarch® PCR DNA Cleanup Kit (New England Biolabs); Am, Chromatin IP DNA Purification Kit (Active Motif); Qp, QIAquick PCR Purification Kit (Qiagen); Qm, MinElute PCR Purification Kit (Qiagen); Ba, Agencourt AMPure XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); Br, RNAClean™ XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); PC, phenol/chloroform extraction. b Interference of PCR amplification by purified eluent of purification reagents. 9 μL eluent was mixed with 1 μL 166 bp of Drosophila probe DNA (0.0001 ng), and the resulting mixture was used as the template in 20 μl of real-time PCR reaction. The Ct value for Drosophila probe DNA from TE buffer was set as 100%. The experiment was repeated 3 times using de-crosslinked chromatin estimated to include 1 ng of DNA. c Size profiles of DNA purified by different reagents. The DNAs purified from de-crosslinked chromatin estimated to include 50 ng range DNA was analyzed by AATI Fragment Analyzer. DNA size (bp) is shown

    Techniques Used: DNA Purification, Chromatin Immunoprecipitation, Purification, Generated, Derivative Assay, Polymerase Chain Reaction, Amplification, Real-time Polymerase Chain Reaction

    2) Product Images from "Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application"

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application

    Journal: BMC Genomics

    doi: 10.1186/s12864-017-4371-5

    DNA purification reagents vary in their ability to recover low amounts of DNA from de-crosslinked chromatin. a Recovered DNA amount by different DNA purification reagents from de-crosslinked chromatin. De-crosslinked chromatin estimated to include 1 ng range DNA in ChIP elution buffer was purified following the manufacturer’s instructions. The data were generated from triplicate DNA samples derived from three independent preparations. Zy, ChIP DNA Clean Concentrator™ (Zymo Research); Pr, Wizard® SV Gel and PCR Clean-Up System (Promega); Th, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, PureLink® PCR Purification Kit (Invitrogen); Ne, Monarch® PCR DNA Cleanup Kit (New England Biolabs); Am, Chromatin IP DNA Purification Kit (Active Motif); Qp, QIAquick PCR Purification Kit (Qiagen); Qm, MinElute PCR Purification Kit (Qiagen); Ba, Agencourt AMPure XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); Br, RNAClean™ XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); PC, phenol/chloroform extraction. b Interference of PCR amplification by purified eluent of purification reagents. 9 μL eluent was mixed with 1 μL 166 bp of Drosophila probe DNA (0.0001 ng), and the resulting mixture was used as the template in 20 μl of real-time PCR reaction. The Ct value for Drosophila probe DNA from TE buffer was set as 100%. The experiment was repeated 3 times using de-crosslinked chromatin estimated to include 1 ng of DNA. c Size profiles of DNA purified by different reagents. The DNAs purified from de-crosslinked chromatin estimated to include 50 ng range DNA was analyzed by AATI Fragment Analyzer. DNA size (bp) is shown
    Figure Legend Snippet: DNA purification reagents vary in their ability to recover low amounts of DNA from de-crosslinked chromatin. a Recovered DNA amount by different DNA purification reagents from de-crosslinked chromatin. De-crosslinked chromatin estimated to include 1 ng range DNA in ChIP elution buffer was purified following the manufacturer’s instructions. The data were generated from triplicate DNA samples derived from three independent preparations. Zy, ChIP DNA Clean Concentrator™ (Zymo Research); Pr, Wizard® SV Gel and PCR Clean-Up System (Promega); Th, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, PureLink® PCR Purification Kit (Invitrogen); Ne, Monarch® PCR DNA Cleanup Kit (New England Biolabs); Am, Chromatin IP DNA Purification Kit (Active Motif); Qp, QIAquick PCR Purification Kit (Qiagen); Qm, MinElute PCR Purification Kit (Qiagen); Ba, Agencourt AMPure XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); Br, RNAClean™ XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); PC, phenol/chloroform extraction. b Interference of PCR amplification by purified eluent of purification reagents. 9 μL eluent was mixed with 1 μL 166 bp of Drosophila probe DNA (0.0001 ng), and the resulting mixture was used as the template in 20 μl of real-time PCR reaction. The Ct value for Drosophila probe DNA from TE buffer was set as 100%. The experiment was repeated 3 times using de-crosslinked chromatin estimated to include 1 ng of DNA. c Size profiles of DNA purified by different reagents. The DNAs purified from de-crosslinked chromatin estimated to include 50 ng range DNA was analyzed by AATI Fragment Analyzer. DNA size (bp) is shown

    Techniques Used: DNA Purification, Chromatin Immunoprecipitation, Purification, Generated, Derivative Assay, Polymerase Chain Reaction, Amplification, Real-time Polymerase Chain Reaction

    Related Articles

    Clone Assay:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: After expansion, genomic DNA extracted from the clones (GeneJET Genomic DNA Purification Kit, K0721, Thermo Fisher Scientific) was used in a polymerase chain reaction (PCR, Phusion Hot Start II DNA polymerase, F549, Thermo Fisher Scientific) to amplify the targeted region. .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Amplification:

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Semi-quantitative RT-PCR was performed via incorporation of [32 P] dCTP [ ].

    Article Title: Target discovery screens using pooled shRNA libraries and next-generation sequencing: A model workflow and analytical algorithm
    Article Snippet: Paragraph title: Genomic DNA isolation and shRNA PCR amplification ... Parallel reaction products were pooled and purified using the GeneJET PCR Purification Kit (Thermo Fisher Scientific).

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Article Title: Geographic mosaic of symbiont selectivity in a genus of epiphytic cyanolichens
    Article Snippet: Special care was taken to ensure that both sequences came from the same lichen thallus and whenever possible both sequences were amplified from the same DNA extraction. .. Amplification products were purified with the GeneJET PCR Purification Kit (Fermentas). .. The tRNALeu (UAA) intron was sequenced with primer pair tRNALeu inF (Paulsrud and Lindblad ) and tRNALeu UR (Fedrowitz et al. ), or with a modified version of the primer tRNALeu inF (tRNALeu UFII, 5′-GGTAGACGCTACGGACTT-3′).

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: DNA templates were added to a reaction mixture containing 1.25 μl of each primer (5 pmoles), 2.5 μl buffer (Bio Basic Inc., Canada), 2 μl MgSO4 (Bio Basic Inc., Canada), 0.3 mM dNTPs, and 0.04 units/μl of taq polymerase (Bio Basic Inc., Canada). .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ). .. SNP genotyping was done by restriction fragment length polymorphism (RFLP) and confirmed by sequencing.

    Article Title: Gut Microbiome Associates With Lipid-Lowering Effect of Rosuvastatin in Vivo
    Article Snippet: After an initial denaturation step at 98°C for 1 min, PCR amplification was carried out using 30 cycles of 98°C for 10 s, 50°C for 30 s and 72°C for 30 s, and a final extension at 72°C for 5 min. PCR reactions were performed on an ABI GeneAmp 9700 PCR system (Applied Biosystems, Foster City, CA, United States). .. The 16S amplicons were purified with GeneJET PCR Purification Kit (Thermo Fisher Scientific, Waltham, MA, United States) and DNA library was constructed by using TruSeq DNA PCR-Free Library Kit (Illumina Inc., San Diego, CA, United States).

    Article Title: An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I
    Article Snippet: The efficiency and sensitivity of the optimized assay was then assessed by analyzing a seven-level standard curve dilution created from purified qPCR product. .. Bull trout ITSI was amplified with the assay and purified using the GeneJET PCR Purification Kit (ThermoFisher Scientific, Waltham, MA, USA). .. The purified product was then quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA), and serially diluted into sterile TE to create a seven-level standard curve (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per reaction).

    Article Title: Prevalence of Genetic Determinants and Phenotypic Resistance to Ciprofloxacin in Campylobacter jejuni from Lithuania
    Article Snippet: The amplification protocol consisted of 95°C for 10 min, followed by 35 cycles of 95°C 30 s, 55°C 30 s, and 72°C 50 s. The reaction was completed by a final extension of 5 min at 72°C. .. The PCR amplicons were purified using the GeneJet PCR purification system (Thermo Scientific, EU).

    Filtration:

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: Parasite DNA was extracted by first removing mouse white blood cells through successive filtration of infected blood using Plasmodipur filters, (Euro-Diagnostica). .. PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer.

    Synthesized:

    Article Title: De Novo Assembly and Characterization of Four Anthozoan (Phylum Cnidaria) Transcriptomes
    Article Snippet: First strand cDNA was synthesized using ProtoScript M-MuLV FS-cDNA Synthesis Kit (New England Biolabs, MA) according to the manufacturer’s protocol and modified oligonucleotides in Supporting Information , Table S1 . .. Second strand synthesis was performed by incubating first-strand cDNA with 1× NEBNext Second Strand Synthesis Buffer (New England Biolabs, MA), 0.2 mM dNTPs, 15 units of Escherichia coli DNA ligase (New England Biolabs, MA), 75 units of E. coli DNA polymerase I (New England Biolabs, MA), and 3 units of RNase H (New England Biolabs, MA) for 2 hr at 16°. cDNA was purified using the GeneJet PCR Purification Kit (Fermentas, MA) and then fragmented using NEBNext dsDNA Fragmentase (New England Biolabs, MA) according to the manufacturer’s protocol, with the addition of 5 mM MgCl2 and 1 mg ml−1 BSA (New England Biolabs, MA).

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: The TK1-TK2 vector was linearized by FseI digest and used to make targeted ES cells for blastocytes injection by standard transgenic mouse techniques at the University of Utah Knockout/Transgenic Mouse Core. .. Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Semi-quantitative RT-PCR was performed via incorporation of [32 P] dCTP [ ].

    Cytometry:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: Two days after transfection, single GFP-positive cells were sorted into separate wells for expansion (MoFlo XDP, Beckman Coulter, CCHMC Research Flow Cytometry Core). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Quantitative RT-PCR:

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas).

    SYBR Green Assay:

    Article Title: Genome-wide mapping of infection-induced SINE RNAs reveals a role in selective mRNA export
    Article Snippet: Cross-linked chromatin was immunoprecipitated using anti-POLR3A (clone ab96328; Abcam) and normal rabbit IgG (clone 2729; Cell Signaling). .. Following reversal of cross-links, DNA was purified using a PCR purification spin column (Fermentas) and resuspended in 50 μl of dH2 O; 1 to 2 μl of DNA was used for quantitative PCR (qPCR) with the DyNAmo ColorFlash SYBR green qPCR kit (Thermo Scientific) with appropriate primers ( ). .. Signals obtained by qPCR were normalized to the input DNA.

    Incubation:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: RNA and protein digestions were performed by addition of 1 μl of 10 μg/μl RNase A (Roche) and incubation at RT for 25 min followed by 11 μl of 10× proteinase K buffer (100 mM Tris-HCl [pH 8.0], 500 mM NaCl, 50 mM EDTA) and 1 μg proteinase K (IBI Scientific) for 1 h at 55°C. .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122).

    Article Title: De Novo Assembly and Characterization of Four Anthozoan (Phylum Cnidaria) Transcriptomes
    Article Snippet: Second strand synthesis was performed by incubating first-strand cDNA with 1× NEBNext Second Strand Synthesis Buffer (New England Biolabs, MA), 0.2 mM dNTPs, 15 units of Escherichia coli DNA ligase (New England Biolabs, MA), 75 units of E. coli DNA polymerase I (New England Biolabs, MA), and 3 units of RNase H (New England Biolabs, MA) for 2 hr at 16°. cDNA was purified using the GeneJet PCR Purification Kit (Fermentas, MA) and then fragmented using NEBNext dsDNA Fragmentase (New England Biolabs, MA) according to the manufacturer’s protocol, with the addition of 5 mM MgCl2 and 1 mg ml−1 BSA (New England Biolabs, MA). .. Tailed templates were ligated to double stranded adaptors prepared with oligonucleotides from the Illumina Customer Sequence Letter (version August 12, 2014, ; Table S1 ).

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: Specifically, 15 μg of DNA was incubated with 640 μM S-adenosylmethionine (SAM) and M. SssI enzyme (5 μL of 20 U/μL) in the provided buffer for 4 h at 37 °C. .. The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water.

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: DNA templates were added to a reaction mixture containing 1.25 μl of each primer (5 pmoles), 2.5 μl buffer (Bio Basic Inc., Canada), 2 μl MgSO4 (Bio Basic Inc., Canada), 0.3 mM dNTPs, and 0.04 units/μl of taq polymerase (Bio Basic Inc., Canada). .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ). .. SNP genotyping was done by restriction fragment length polymorphism (RFLP) and confirmed by sequencing.

    Expressing:

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Products were visualized by exposure to X-ray film at -80° C. Quantitative RT-PCR was performed using Light Cycler (Roche Diagnostics) [ ].

    Modification:

    Article Title: De Novo Assembly and Characterization of Four Anthozoan (Phylum Cnidaria) Transcriptomes
    Article Snippet: First strand cDNA was synthesized using ProtoScript M-MuLV FS-cDNA Synthesis Kit (New England Biolabs, MA) according to the manufacturer’s protocol and modified oligonucleotides in Supporting Information , Table S1 . .. Second strand synthesis was performed by incubating first-strand cDNA with 1× NEBNext Second Strand Synthesis Buffer (New England Biolabs, MA), 0.2 mM dNTPs, 15 units of Escherichia coli DNA ligase (New England Biolabs, MA), 75 units of E. coli DNA polymerase I (New England Biolabs, MA), and 3 units of RNase H (New England Biolabs, MA) for 2 hr at 16°. cDNA was purified using the GeneJet PCR Purification Kit (Fermentas, MA) and then fragmented using NEBNext dsDNA Fragmentase (New England Biolabs, MA) according to the manufacturer’s protocol, with the addition of 5 mM MgCl2 and 1 mg ml−1 BSA (New England Biolabs, MA).

    Western Blot:

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    Article Snippet: The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific). .. A seven‐level serial dilution (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per 4 μl) was created in sterile TE, and each level was analyzed in six replicates.

    DNA Methylation Assay:

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: Paragraph title: Nucleosome Assembly and DNA Methylation ... The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water.

    Derivative Assay:

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ). .. Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ).

    High Performance Liquid Chromatography:

    Article Title: Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine
    Article Snippet: Genomic DNA sample (100 ng–1 μg) Illumina TruSeq DNA sample preparation kit (Illumina, cat. no. FC-121-2001) Ampure XP beads (Beckman Coulter, cat. no. ) Milli-Q water Sodium hydroxide, 1.0 M (NaOH; Sigma-Aldrich, cat. no. 319511) Oxidant solution (10× solution; Cambridge Epigenetix (CEGX)). .. Alternatively, in place of Steps 12–14 and 17, the oxidant and oxidation protocol can be prepared and used as described in (ref. ) Hydroxymethyl dCTP (dhmCTP, Bioline, cat. no. BIO-39046) dNTP mix, 10 mM (NEB, cat. no. N0447S) Epitect bisulfite kit (Qiagen, cat. no. 59104) P-6 saline–sodium citrate (SSC) Micro Bio-Spin columns (Bio-Rad, cat. no. 732-6200) Pfu Turbo Cx hotstart DNA polymerase (Agilent, cat. no. 600410) Agarose (Sigma-Aldrich, cat. no. A9539) dATP, 100 mM (NEB, cat. no. N0446S) dGTP, 100 mM (NEB, cat. no. N0446S) dTTP, 100 mM (NEB, cat. no. N0446S) DNA templates (Biomers; ) DNA primers (Biomers; ) DreamTaq DNA polymerase (Thermo Scientific, cat. no. EP0701) KAPA HiFi uracil + ReadyMix (KAPA Biosystems, cat. no. KK2801) GeneJet PCR purification kit (Thermo Scientific, cat. no. K0701) Taqa1 restriction endonuclease (Taq1 RE; NEB, cat. no. R0149S) Ethanol (Sigma-Aldrich, cat. no. 32221) GelRed, 3× dye (Cambridge Bioscience, cat. no. 41001) Ammonium acetate (Fisher, cat. no. A/3440/53) Acetic acid (Sigma-Aldrich, cat. no. 320099) HPLC-grade acetonitrile (Sigma-Aldrich, cat. no. 34851) .. Manual pipettes (Thermo Finnpipettes F1, cat. nos.

    Flow Cytometry:

    Article Title: Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine
    Article Snippet: Genomic DNA sample (100 ng–1 μg) Illumina TruSeq DNA sample preparation kit (Illumina, cat. no. FC-121-2001) Ampure XP beads (Beckman Coulter, cat. no. ) Milli-Q water Sodium hydroxide, 1.0 M (NaOH; Sigma-Aldrich, cat. no. 319511) Oxidant solution (10× solution; Cambridge Epigenetix (CEGX)). .. Alternatively, in place of Steps 12–14 and 17, the oxidant and oxidation protocol can be prepared and used as described in (ref. ) Hydroxymethyl dCTP (dhmCTP, Bioline, cat. no. BIO-39046) dNTP mix, 10 mM (NEB, cat. no. N0447S) Epitect bisulfite kit (Qiagen, cat. no. 59104) P-6 saline–sodium citrate (SSC) Micro Bio-Spin columns (Bio-Rad, cat. no. 732-6200) Pfu Turbo Cx hotstart DNA polymerase (Agilent, cat. no. 600410) Agarose (Sigma-Aldrich, cat. no. A9539) dATP, 100 mM (NEB, cat. no. N0446S) dGTP, 100 mM (NEB, cat. no. N0446S) dTTP, 100 mM (NEB, cat. no. N0446S) DNA templates (Biomers; ) DNA primers (Biomers; ) DreamTaq DNA polymerase (Thermo Scientific, cat. no. EP0701) KAPA HiFi uracil + ReadyMix (KAPA Biosystems, cat. no. KK2801) GeneJet PCR purification kit (Thermo Scientific, cat. no. K0701) Taqa1 restriction endonuclease (Taq1 RE; NEB, cat. no. R0149S) Ethanol (Sigma-Aldrich, cat. no. 32221) GelRed, 3× dye (Cambridge Bioscience, cat. no. 41001) Ammonium acetate (Fisher, cat. no. A/3440/53) Acetic acid (Sigma-Aldrich, cat. no. 320099) HPLC-grade acetonitrile (Sigma-Aldrich, cat. no. 34851)

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: Two days after transfection, single GFP-positive cells were sorted into separate wells for expansion (MoFlo XDP, Beckman Coulter, CCHMC Research Flow Cytometry Core). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Gas Chromatography:

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: The primers (Integrated DNA Technology, USA) were sized between 22 and 24 bases with a Tm of 69–71 °C and a GC content of 40–60%. .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ).

    Sensitive Assay:

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: DNA was purified using MinElute PCR Purification Kit after treatment of RNase A and proteinase K. DNA was quantified using Qubit dsDNA High Sensitivity assay and adjusted to 1 ng/μL with TE buffer. .. Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ).

    Ligation:

    Article Title: Synthetic Peptides to Target Stringent Response-Controlled Virulence in a Pseudomonas aeruginosa Murine Cutaneous Infection Model
    Article Snippet: All ligation reactions were carried out at room temperature using Thermo Scientific T4 DNA ligase. .. DNA purifications were either performed using the GeneJET PCR purification kit (Thermo Scientific) or the GeneJET Gel extraction kit (Thermo Scientific) following the manufacturer’s instructions.

    Infection:

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: Parasite DNA was extracted by first removing mouse white blood cells through successive filtration of infected blood using Plasmodipur filters, (Euro-Diagnostica). .. PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer.

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: Paragraph title: RNA preparation, cDNA synthesis and RT-PCR ... Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas).

    Irradiation:

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: Experiments were set up inside a hood where qPCR consumables and pipettes were irradiated with UV for 1 h before setup. .. The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific).

    Generated:

    Article Title: Prevalence of Genetic Determinants and Phenotypic Resistance to Ciprofloxacin in Campylobacter jejuni from Lithuania
    Article Snippet: The partial gene sequence of gyrA , generated by use of CampyMAMAgyrA1 and a reverse primer CampyMAMAgyrA5 targeting the quinolone resistance determining region (QRDR), was sequenced with the forward and reverse primers described by Ragimbeau et al. (Table ). .. The PCR amplicons were purified using the GeneJet PCR purification system (Thermo Scientific, EU).

    Imaging:

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: Gel shift was completed by running 10 μL of the assembly reaction on a 6% polyacrylamide gel, staining the DNA with SYBR-safe (Life Technologies S33102), and imaging with a Chemi-doc imager (Biorad) ( Figure S4). .. The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water.

    Polymerase Chain Reaction:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: RNA and protein digestions were performed by addition of 1 μl of 10 μg/μl RNase A (Roche) and incubation at RT for 25 min followed by 11 μl of 10× proteinase K buffer (100 mM Tris-HCl [pH 8.0], 500 mM NaCl, 50 mM EDTA) and 1 μg proteinase K (IBI Scientific) for 1 h at 55°C. .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122). .. Libraries were submitted for 50-bp single-end Illumina sequencing as described above.

    Article Title: De Novo Assembly and Characterization of Four Anthozoan (Phylum Cnidaria) Transcriptomes
    Article Snippet: First strand cDNA was synthesized using ProtoScript M-MuLV FS-cDNA Synthesis Kit (New England Biolabs, MA) according to the manufacturer’s protocol and modified oligonucleotides in Supporting Information , Table S1 . .. Second strand synthesis was performed by incubating first-strand cDNA with 1× NEBNext Second Strand Synthesis Buffer (New England Biolabs, MA), 0.2 mM dNTPs, 15 units of Escherichia coli DNA ligase (New England Biolabs, MA), 75 units of E. coli DNA polymerase I (New England Biolabs, MA), and 3 units of RNase H (New England Biolabs, MA) for 2 hr at 16°. cDNA was purified using the GeneJet PCR Purification Kit (Fermentas, MA) and then fragmented using NEBNext dsDNA Fragmentase (New England Biolabs, MA) according to the manufacturer’s protocol, with the addition of 5 mM MgCl2 and 1 mg ml−1 BSA (New England Biolabs, MA). .. Fragmented cDNA was purified and the ends were repaired using NEB Quick Blunting Kit (New England Biolabs, MA) according to manufacturer’s protocol.

    Article Title: Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine
    Article Snippet: Genomic DNA sample (100 ng–1 μg) Illumina TruSeq DNA sample preparation kit (Illumina, cat. no. FC-121-2001) Ampure XP beads (Beckman Coulter, cat. no. ) Milli-Q water Sodium hydroxide, 1.0 M (NaOH; Sigma-Aldrich, cat. no. 319511) Oxidant solution (10× solution; Cambridge Epigenetix (CEGX)). .. Alternatively, in place of Steps 12–14 and 17, the oxidant and oxidation protocol can be prepared and used as described in (ref. ) Hydroxymethyl dCTP (dhmCTP, Bioline, cat. no. BIO-39046) dNTP mix, 10 mM (NEB, cat. no. N0447S) Epitect bisulfite kit (Qiagen, cat. no. 59104) P-6 saline–sodium citrate (SSC) Micro Bio-Spin columns (Bio-Rad, cat. no. 732-6200) Pfu Turbo Cx hotstart DNA polymerase (Agilent, cat. no. 600410) Agarose (Sigma-Aldrich, cat. no. A9539) dATP, 100 mM (NEB, cat. no. N0446S) dGTP, 100 mM (NEB, cat. no. N0446S) dTTP, 100 mM (NEB, cat. no. N0446S) DNA templates (Biomers; ) DNA primers (Biomers; ) DreamTaq DNA polymerase (Thermo Scientific, cat. no. EP0701) KAPA HiFi uracil + ReadyMix (KAPA Biosystems, cat. no. KK2801) GeneJet PCR purification kit (Thermo Scientific, cat. no. K0701) Taqa1 restriction endonuclease (Taq1 RE; NEB, cat. no. R0149S) Ethanol (Sigma-Aldrich, cat. no. 32221) GelRed, 3× dye (Cambridge Bioscience, cat. no. 41001) Ammonium acetate (Fisher, cat. no. A/3440/53) Acetic acid (Sigma-Aldrich, cat. no. 320099) HPLC-grade acetonitrile (Sigma-Aldrich, cat. no. 34851) .. Manual pipettes (Thermo Finnpipettes F1, cat. nos.

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: The TK1-TK2 vector was linearized by FseI digest and used to make targeted ES cells for blastocytes injection by standard transgenic mouse techniques at the University of Utah Knockout/Transgenic Mouse Core. .. Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Semi-quantitative RT-PCR was performed via incorporation of [32 P] dCTP [ ].

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: However, the yields of DNA recovery varied considerably among the kits. .. The Wizard® SV Gel and PCR Clean-Up System (Promega; Pr), the GeneJET PCR Purification Kit (Thermo Fisher Scientific; Th), the PureLink® PCR Purification Kit (Invitrogen; In) and the Chromatin IP DNA Purification Kit (Active Motif; Am) performed poorly with de-crosslinked chromatin. .. Consistently, these reagents recovered less than 50% of input DNA with purified DNA even when expected DNA amounts were relatively high (10–50 ng).

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: DNAs were prepared to final 1 ng, 5 ng, 10 ng and 50 ng in 100 μL ChIP elution buffer and were purified by 11 different purification reagents as suggested by the manufacturer except for the elution volume, which was fixed at 16 μL. .. Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ). .. The sample-to-beads ratio tested for Ba and Br were 1:1.25, 1:1.50, 1:1.75, and 1:2.

    Article Title: Target discovery screens using pooled shRNA libraries and next-generation sequencing: A model workflow and analytical algorithm
    Article Snippet: PCR was performed at 98°C for 1.5 min followed by 33 cycles of 98°C (10 s), 56°C (30 s), 72°C (30 s), and final 72°C (10min) on a TProfessional cycler (Biometra, Göttingen, Germany). .. Parallel reaction products were pooled and purified using the GeneJET PCR Purification Kit (Thermo Fisher Scientific). .. Product quantity and quality were evaluated using NanoDrop spectrophotometry, Qubit fluorometric quantification (Qubit 3.0 and dsDNA high-sensitivity assay; Thermo Fisher Scientific) and Bioanalyzer gel electrophoresis (Bioanalyzer 2100 and Agilent DNA 1000 Kit; Agilent Technologies, Santa Clara, CA).

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: Specifically, 15 μg of DNA was incubated with 640 μM S-adenosylmethionine (SAM) and M. SssI enzyme (5 μL of 20 U/μL) in the provided buffer for 4 h at 37 °C. .. The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water. .. DNA was quantified, lyophilized, and resuspended at the appropriate concentration (10 μM).

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: These PCR products were screened for loss of the AflIII restriction site (New England Biolabs, Ipswich, MA). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core). .. We selected clonal lines that had, in both alleles, a frameshifting mutation that resulted in an early (first exon) stop codon.

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: The other reagents MgCl2 , dNTPs, forward and reverse primers, Dream Taq Polymerase (Thermo-Scientific) and cycling conditions were optimized accordingly as shown in . .. PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer. .. Contigs were assembled using Lasergene 11 Core Suite, the DNA sequences and the predicted amino acid sequences were analysed using CLUSTAL W available in EBI website ( ) and searched by BLAST© software available at the NCBI Website and PlasmoDB version 11.0 ( ).

    Article Title: Genotyping of Plant and Animal Samples without Prior DNA Purification
    Article Snippet: When amplifying DNA directly from plant tissues, the PCR products include plant and PCR-derived components that may interfere with the restriction digestion enzyme. .. Therefore, it may be necessary to either dilute ( e.g. 1:2 or 1:3 in water) or purify the PCR product before the subsequent digestion, for example by using a suitable commercial kit such as Thermo Scientific GeneJET PCR Purification Kit. .. Following restriction enzyme digestion, analyze the resulting fragments on an agarose gel.

    Article Title: Synthetic Peptides to Target Stringent Response-Controlled Virulence in a Pseudomonas aeruginosa Murine Cutaneous Infection Model
    Article Snippet: All ligation reactions were carried out at room temperature using Thermo Scientific T4 DNA ligase. .. DNA purifications were either performed using the GeneJET PCR purification kit (Thermo Scientific) or the GeneJET Gel extraction kit (Thermo Scientific) following the manufacturer’s instructions. .. Peptides 1018 (VRLIVAVRIWRR-NH2 ) and DJK-5 (VQWRAIRVRVIR-NH2 ) were synthesized by CPC Scientific using solid-phase 9-flurenylmethoxy carbonyl (Fmoc) chemistry and purified to > 95% purity using reverse-phase high-performance liquid chromatography (HPLC).

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: Using these optimized primer concentrations, we then performed a standard curve analysis to examine the sensitivity of the assay. .. The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific). .. A seven‐level serial dilution (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per 4 μl) was created in sterile TE, and each level was analyzed in six replicates.

    Article Title: Geographic mosaic of symbiont selectivity in a genus of epiphytic cyanolichens
    Article Snippet: Special care was taken to ensure that both sequences came from the same lichen thallus and whenever possible both sequences were amplified from the same DNA extraction. .. Amplification products were purified with the GeneJET PCR Purification Kit (Fermentas). .. The tRNALeu (UAA) intron was sequenced with primer pair tRNALeu inF (Paulsrud and Lindblad ) and tRNALeu UR (Fedrowitz et al. ), or with a modified version of the primer tRNALeu inF (tRNALeu UFII, 5′-GGTAGACGCTACGGACTT-3′).

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: DNA templates were added to a reaction mixture containing 1.25 μl of each primer (5 pmoles), 2.5 μl buffer (Bio Basic Inc., Canada), 2 μl MgSO4 (Bio Basic Inc., Canada), 0.3 mM dNTPs, and 0.04 units/μl of taq polymerase (Bio Basic Inc., Canada). .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ). .. SNP genotyping was done by restriction fragment length polymorphism (RFLP) and confirmed by sequencing.

    Article Title: Gut Microbiome Associates With Lipid-Lowering Effect of Rosuvastatin in Vivo
    Article Snippet: After an initial denaturation step at 98°C for 1 min, PCR amplification was carried out using 30 cycles of 98°C for 10 s, 50°C for 30 s and 72°C for 30 s, and a final extension at 72°C for 5 min. PCR reactions were performed on an ABI GeneAmp 9700 PCR system (Applied Biosystems, Foster City, CA, United States). .. The 16S amplicons were purified with GeneJET PCR Purification Kit (Thermo Fisher Scientific, Waltham, MA, United States) and DNA library was constructed by using TruSeq DNA PCR-Free Library Kit (Illumina Inc., San Diego, CA, United States). .. The libraries were then sequenced by Hiseq sequencing System (Illumina Inc., San Diego, CA, United States).

    Article Title: An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I
    Article Snippet: The efficiency and sensitivity of the optimized assay was then assessed by analyzing a seven-level standard curve dilution created from purified qPCR product. .. Bull trout ITSI was amplified with the assay and purified using the GeneJET PCR Purification Kit (ThermoFisher Scientific, Waltham, MA, USA). .. The purified product was then quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA), and serially diluted into sterile TE to create a seven-level standard curve (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per reaction).

    Article Title: Prevalence of Genetic Determinants and Phenotypic Resistance to Ciprofloxacin in Campylobacter jejuni from Lithuania
    Article Snippet: The amplification protocol consisted of 95°C for 10 min, followed by 35 cycles of 95°C 30 s, 55°C 30 s, and 72°C 50 s. The reaction was completed by a final extension of 5 min at 72°C. .. The PCR amplicons were purified using the GeneJet PCR purification system (Thermo Scientific, EU). .. Sequencing reactions were carried out using the BigDye Terminator 3.1 cycle Sequencing Kit (Applied Biosystems, USA) according to instructions from the manufacturer.

    Article Title: Genome-wide mapping of infection-induced SINE RNAs reveals a role in selective mRNA export
    Article Snippet: Cross-linked chromatin was immunoprecipitated using anti-POLR3A (clone ab96328; Abcam) and normal rabbit IgG (clone 2729; Cell Signaling). .. Following reversal of cross-links, DNA was purified using a PCR purification spin column (Fermentas) and resuspended in 50 μl of dH2 O; 1 to 2 μl of DNA was used for quantitative PCR (qPCR) with the DyNAmo ColorFlash SYBR green qPCR kit (Thermo Scientific) with appropriate primers ( ). .. Signals obtained by qPCR were normalized to the input DNA.

    Molecular Weight:

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: DNA templates were added to a reaction mixture containing 1.25 μl of each primer (5 pmoles), 2.5 μl buffer (Bio Basic Inc., Canada), 2 μl MgSO4 (Bio Basic Inc., Canada), 0.3 mM dNTPs, and 0.04 units/μl of taq polymerase (Bio Basic Inc., Canada). .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ). .. SNP genotyping was done by restriction fragment length polymorphism (RFLP) and confirmed by sequencing.

    DNA Sequencing:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: These PCR products were screened for loss of the AflIII restriction site (New England Biolabs, Ipswich, MA). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core). .. We selected clonal lines that had, in both alleles, a frameshifting mutation that resulted in an early (first exon) stop codon.

    Article Title: Prevalence of Genetic Determinants and Phenotypic Resistance to Ciprofloxacin in Campylobacter jejuni from Lithuania
    Article Snippet: Paragraph title: DNA sequencing and sequence analysis ... The PCR amplicons were purified using the GeneJet PCR purification system (Thermo Scientific, EU).

    DNA Extraction:

    Article Title: Target discovery screens using pooled shRNA libraries and next-generation sequencing: A model workflow and analytical algorithm
    Article Snippet: Paragraph title: Genomic DNA isolation and shRNA PCR amplification ... Parallel reaction products were pooled and purified using the GeneJET PCR Purification Kit (Thermo Fisher Scientific).

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: Paragraph title: 2.5. DNA extraction, PCR and Sequencing ... PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer.

    Article Title: Geographic mosaic of symbiont selectivity in a genus of epiphytic cyanolichens
    Article Snippet: Paragraph title: DNA extraction, amplification, and sequencing ... Amplification products were purified with the GeneJET PCR Purification Kit (Fermentas).

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: Paragraph title: DNA Extraction and Amplification ... The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ).

    Nucleic Acid Electrophoresis:

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Semi-quantitative RT-PCR was performed via incorporation of [32 P] dCTP [ ].

    In Vivo:

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific). .. A seven‐level serial dilution (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per 4 μl) was created in sterile TE, and each level was analyzed in six replicates.

    Gene Knockout:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    ChIP-sequencing:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: RNA and protein digestions were performed by addition of 1 μl of 10 μg/μl RNase A (Roche) and incubation at RT for 25 min followed by 11 μl of 10× proteinase K buffer (100 mM Tris-HCl [pH 8.0], 500 mM NaCl, 50 mM EDTA) and 1 μg proteinase K (IBI Scientific) for 1 h at 55°C. .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122). .. Libraries were submitted for 50-bp single-end Illumina sequencing as described above.

    Fluorescence:

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific). .. The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific).

    Article Title: An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I
    Article Snippet: The primer concentrations producing the earliest cycle threshold (Ct) value and highest end-point fluorescence were selected for further analysis. .. Bull trout ITSI was amplified with the assay and purified using the GeneJET PCR Purification Kit (ThermoFisher Scientific, Waltham, MA, USA).

    Methylation:

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: DNA was methylated using CpG Methyltransferase (M.SssI) (NEB M0226M). .. The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water.

    Mutagenesis:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: The Core validated the mutation efficiency in a murine cell line with a T7 endonuclease I test. .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Isolation:

    Article Title: De Novo Assembly and Characterization of Four Anthozoan (Phylum Cnidaria) Transcriptomes
    Article Snippet: Polyadenylated RNA was purified from 10 µg of total RNA using the Magnetic mRNA Isolation Kit (New England Biolabs, MA). .. Second strand synthesis was performed by incubating first-strand cDNA with 1× NEBNext Second Strand Synthesis Buffer (New England Biolabs, MA), 0.2 mM dNTPs, 15 units of Escherichia coli DNA ligase (New England Biolabs, MA), 75 units of E. coli DNA polymerase I (New England Biolabs, MA), and 3 units of RNase H (New England Biolabs, MA) for 2 hr at 16°. cDNA was purified using the GeneJet PCR Purification Kit (Fermentas, MA) and then fragmented using NEBNext dsDNA Fragmentase (New England Biolabs, MA) according to the manufacturer’s protocol, with the addition of 5 mM MgCl2 and 1 mg ml−1 BSA (New England Biolabs, MA).

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: The TK1-TK2 vector was linearized by FseI digest and used to make targeted ES cells for blastocytes injection by standard transgenic mouse techniques at the University of Utah Knockout/Transgenic Mouse Core. .. Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Semi-quantitative RT-PCR was performed via incorporation of [32 P] dCTP [ ].

    Article Title: Target discovery screens using pooled shRNA libraries and next-generation sequencing: A model workflow and analytical algorithm
    Article Snippet: Parallel reaction products were pooled and purified using the GeneJET PCR Purification Kit (Thermo Fisher Scientific). .. Product quantity and quality were evaluated using NanoDrop spectrophotometry, Qubit fluorometric quantification (Qubit 3.0 and dsDNA high-sensitivity assay; Thermo Fisher Scientific) and Bioanalyzer gel electrophoresis (Bioanalyzer 2100 and Agilent DNA 1000 Kit; Agilent Technologies, Santa Clara, CA).

    Transfection:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: Two days after transfection, single GFP-positive cells were sorted into separate wells for expansion (MoFlo XDP, Beckman Coulter, CCHMC Research Flow Cytometry Core). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Electrophoretic Mobility Shift Assay:

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: Gel shift was completed by running 10 μL of the assembly reaction on a 6% polyacrylamide gel, staining the DNA with SYBR-safe (Life Technologies S33102), and imaging with a Chemi-doc imager (Biorad) ( Figure S4). .. The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water.

    Purification:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: RNA and protein digestions were performed by addition of 1 μl of 10 μg/μl RNase A (Roche) and incubation at RT for 25 min followed by 11 μl of 10× proteinase K buffer (100 mM Tris-HCl [pH 8.0], 500 mM NaCl, 50 mM EDTA) and 1 μg proteinase K (IBI Scientific) for 1 h at 55°C. .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122). .. Libraries were submitted for 50-bp single-end Illumina sequencing as described above.

    Article Title: De Novo Assembly and Characterization of Four Anthozoan (Phylum Cnidaria) Transcriptomes
    Article Snippet: First strand cDNA was synthesized using ProtoScript M-MuLV FS-cDNA Synthesis Kit (New England Biolabs, MA) according to the manufacturer’s protocol and modified oligonucleotides in Supporting Information , Table S1 . .. Second strand synthesis was performed by incubating first-strand cDNA with 1× NEBNext Second Strand Synthesis Buffer (New England Biolabs, MA), 0.2 mM dNTPs, 15 units of Escherichia coli DNA ligase (New England Biolabs, MA), 75 units of E. coli DNA polymerase I (New England Biolabs, MA), and 3 units of RNase H (New England Biolabs, MA) for 2 hr at 16°. cDNA was purified using the GeneJet PCR Purification Kit (Fermentas, MA) and then fragmented using NEBNext dsDNA Fragmentase (New England Biolabs, MA) according to the manufacturer’s protocol, with the addition of 5 mM MgCl2 and 1 mg ml−1 BSA (New England Biolabs, MA). .. Fragmented cDNA was purified and the ends were repaired using NEB Quick Blunting Kit (New England Biolabs, MA) according to manufacturer’s protocol.

    Article Title: Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine
    Article Snippet: Genomic DNA sample (100 ng–1 μg) Illumina TruSeq DNA sample preparation kit (Illumina, cat. no. FC-121-2001) Ampure XP beads (Beckman Coulter, cat. no. ) Milli-Q water Sodium hydroxide, 1.0 M (NaOH; Sigma-Aldrich, cat. no. 319511) Oxidant solution (10× solution; Cambridge Epigenetix (CEGX)). .. Alternatively, in place of Steps 12–14 and 17, the oxidant and oxidation protocol can be prepared and used as described in (ref. ) Hydroxymethyl dCTP (dhmCTP, Bioline, cat. no. BIO-39046) dNTP mix, 10 mM (NEB, cat. no. N0447S) Epitect bisulfite kit (Qiagen, cat. no. 59104) P-6 saline–sodium citrate (SSC) Micro Bio-Spin columns (Bio-Rad, cat. no. 732-6200) Pfu Turbo Cx hotstart DNA polymerase (Agilent, cat. no. 600410) Agarose (Sigma-Aldrich, cat. no. A9539) dATP, 100 mM (NEB, cat. no. N0446S) dGTP, 100 mM (NEB, cat. no. N0446S) dTTP, 100 mM (NEB, cat. no. N0446S) DNA templates (Biomers; ) DNA primers (Biomers; ) DreamTaq DNA polymerase (Thermo Scientific, cat. no. EP0701) KAPA HiFi uracil + ReadyMix (KAPA Biosystems, cat. no. KK2801) GeneJet PCR purification kit (Thermo Scientific, cat. no. K0701) Taqa1 restriction endonuclease (Taq1 RE; NEB, cat. no. R0149S) Ethanol (Sigma-Aldrich, cat. no. 32221) GelRed, 3× dye (Cambridge Bioscience, cat. no. 41001) Ammonium acetate (Fisher, cat. no. A/3440/53) Acetic acid (Sigma-Aldrich, cat. no. 320099) HPLC-grade acetonitrile (Sigma-Aldrich, cat. no. 34851) .. Manual pipettes (Thermo Finnpipettes F1, cat. nos.

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: The TK1-TK2 vector was linearized by FseI digest and used to make targeted ES cells for blastocytes injection by standard transgenic mouse techniques at the University of Utah Knockout/Transgenic Mouse Core. .. Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Semi-quantitative RT-PCR was performed via incorporation of [32 P] dCTP [ ].

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: However, the yields of DNA recovery varied considerably among the kits. .. The Wizard® SV Gel and PCR Clean-Up System (Promega; Pr), the GeneJET PCR Purification Kit (Thermo Fisher Scientific; Th), the PureLink® PCR Purification Kit (Invitrogen; In) and the Chromatin IP DNA Purification Kit (Active Motif; Am) performed poorly with de-crosslinked chromatin. .. Consistently, these reagents recovered less than 50% of input DNA with purified DNA even when expected DNA amounts were relatively high (10–50 ng).

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: DNAs were prepared to final 1 ng, 5 ng, 10 ng and 50 ng in 100 μL ChIP elution buffer and were purified by 11 different purification reagents as suggested by the manufacturer except for the elution volume, which was fixed at 16 μL. .. Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ). .. The sample-to-beads ratio tested for Ba and Br were 1:1.25, 1:1.50, 1:1.75, and 1:2.

    Article Title: Target discovery screens using pooled shRNA libraries and next-generation sequencing: A model workflow and analytical algorithm
    Article Snippet: PCR was performed at 98°C for 1.5 min followed by 33 cycles of 98°C (10 s), 56°C (30 s), 72°C (30 s), and final 72°C (10min) on a TProfessional cycler (Biometra, Göttingen, Germany). .. Parallel reaction products were pooled and purified using the GeneJET PCR Purification Kit (Thermo Fisher Scientific). .. Product quantity and quality were evaluated using NanoDrop spectrophotometry, Qubit fluorometric quantification (Qubit 3.0 and dsDNA high-sensitivity assay; Thermo Fisher Scientific) and Bioanalyzer gel electrophoresis (Bioanalyzer 2100 and Agilent DNA 1000 Kit; Agilent Technologies, Santa Clara, CA).

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: Specifically, 15 μg of DNA was incubated with 640 μM S-adenosylmethionine (SAM) and M. SssI enzyme (5 μL of 20 U/μL) in the provided buffer for 4 h at 37 °C. .. The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water. .. DNA was quantified, lyophilized, and resuspended at the appropriate concentration (10 μM).

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: These PCR products were screened for loss of the AflIII restriction site (New England Biolabs, Ipswich, MA). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core). .. We selected clonal lines that had, in both alleles, a frameshifting mutation that resulted in an early (first exon) stop codon.

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: The other reagents MgCl2 , dNTPs, forward and reverse primers, Dream Taq Polymerase (Thermo-Scientific) and cycling conditions were optimized accordingly as shown in . .. PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer. .. Contigs were assembled using Lasergene 11 Core Suite, the DNA sequences and the predicted amino acid sequences were analysed using CLUSTAL W available in EBI website ( ) and searched by BLAST© software available at the NCBI Website and PlasmoDB version 11.0 ( ).

    Article Title: Genotyping of Plant and Animal Samples without Prior DNA Purification
    Article Snippet: When amplifying DNA directly from plant tissues, the PCR products include plant and PCR-derived components that may interfere with the restriction digestion enzyme. .. Therefore, it may be necessary to either dilute ( e.g. 1:2 or 1:3 in water) or purify the PCR product before the subsequent digestion, for example by using a suitable commercial kit such as Thermo Scientific GeneJET PCR Purification Kit. .. Following restriction enzyme digestion, analyze the resulting fragments on an agarose gel.

    Article Title: Synthetic Peptides to Target Stringent Response-Controlled Virulence in a Pseudomonas aeruginosa Murine Cutaneous Infection Model
    Article Snippet: All ligation reactions were carried out at room temperature using Thermo Scientific T4 DNA ligase. .. DNA purifications were either performed using the GeneJET PCR purification kit (Thermo Scientific) or the GeneJET Gel extraction kit (Thermo Scientific) following the manufacturer’s instructions. .. Peptides 1018 (VRLIVAVRIWRR-NH2 ) and DJK-5 (VQWRAIRVRVIR-NH2 ) were synthesized by CPC Scientific using solid-phase 9-flurenylmethoxy carbonyl (Fmoc) chemistry and purified to > 95% purity using reverse-phase high-performance liquid chromatography (HPLC).

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: Using these optimized primer concentrations, we then performed a standard curve analysis to examine the sensitivity of the assay. .. The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific). .. A seven‐level serial dilution (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per 4 μl) was created in sterile TE, and each level was analyzed in six replicates.

    Article Title: Geographic mosaic of symbiont selectivity in a genus of epiphytic cyanolichens
    Article Snippet: Special care was taken to ensure that both sequences came from the same lichen thallus and whenever possible both sequences were amplified from the same DNA extraction. .. Amplification products were purified with the GeneJET PCR Purification Kit (Fermentas). .. The tRNALeu (UAA) intron was sequenced with primer pair tRNALeu inF (Paulsrud and Lindblad ) and tRNALeu UR (Fedrowitz et al. ), or with a modified version of the primer tRNALeu inF (tRNALeu UFII, 5′-GGTAGACGCTACGGACTT-3′).

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: DNA templates were added to a reaction mixture containing 1.25 μl of each primer (5 pmoles), 2.5 μl buffer (Bio Basic Inc., Canada), 2 μl MgSO4 (Bio Basic Inc., Canada), 0.3 mM dNTPs, and 0.04 units/μl of taq polymerase (Bio Basic Inc., Canada). .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ). .. SNP genotyping was done by restriction fragment length polymorphism (RFLP) and confirmed by sequencing.

    Article Title: Gut Microbiome Associates With Lipid-Lowering Effect of Rosuvastatin in Vivo
    Article Snippet: After an initial denaturation step at 98°C for 1 min, PCR amplification was carried out using 30 cycles of 98°C for 10 s, 50°C for 30 s and 72°C for 30 s, and a final extension at 72°C for 5 min. PCR reactions were performed on an ABI GeneAmp 9700 PCR system (Applied Biosystems, Foster City, CA, United States). .. The 16S amplicons were purified with GeneJET PCR Purification Kit (Thermo Fisher Scientific, Waltham, MA, United States) and DNA library was constructed by using TruSeq DNA PCR-Free Library Kit (Illumina Inc., San Diego, CA, United States). .. The libraries were then sequenced by Hiseq sequencing System (Illumina Inc., San Diego, CA, United States).

    Article Title: An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I
    Article Snippet: The efficiency and sensitivity of the optimized assay was then assessed by analyzing a seven-level standard curve dilution created from purified qPCR product. .. Bull trout ITSI was amplified with the assay and purified using the GeneJET PCR Purification Kit (ThermoFisher Scientific, Waltham, MA, USA). .. The purified product was then quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA), and serially diluted into sterile TE to create a seven-level standard curve (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per reaction).

    Article Title: Prevalence of Genetic Determinants and Phenotypic Resistance to Ciprofloxacin in Campylobacter jejuni from Lithuania
    Article Snippet: The amplification protocol consisted of 95°C for 10 min, followed by 35 cycles of 95°C 30 s, 55°C 30 s, and 72°C 50 s. The reaction was completed by a final extension of 5 min at 72°C. .. The PCR amplicons were purified using the GeneJet PCR purification system (Thermo Scientific, EU). .. Sequencing reactions were carried out using the BigDye Terminator 3.1 cycle Sequencing Kit (Applied Biosystems, USA) according to instructions from the manufacturer.

    Article Title: Genome-wide mapping of infection-induced SINE RNAs reveals a role in selective mRNA export
    Article Snippet: Cross-linked chromatin was immunoprecipitated using anti-POLR3A (clone ab96328; Abcam) and normal rabbit IgG (clone 2729; Cell Signaling). .. Following reversal of cross-links, DNA was purified using a PCR purification spin column (Fermentas) and resuspended in 50 μl of dH2 O; 1 to 2 μl of DNA was used for quantitative PCR (qPCR) with the DyNAmo ColorFlash SYBR green qPCR kit (Thermo Scientific) with appropriate primers ( ). .. Signals obtained by qPCR were normalized to the input DNA.

    Sequencing:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122). .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122).

    Article Title: De Novo Assembly and Characterization of Four Anthozoan (Phylum Cnidaria) Transcriptomes
    Article Snippet: Paragraph title: Preparation of sequencing libraries ... Second strand synthesis was performed by incubating first-strand cDNA with 1× NEBNext Second Strand Synthesis Buffer (New England Biolabs, MA), 0.2 mM dNTPs, 15 units of Escherichia coli DNA ligase (New England Biolabs, MA), 75 units of E. coli DNA polymerase I (New England Biolabs, MA), and 3 units of RNase H (New England Biolabs, MA) for 2 hr at 16°. cDNA was purified using the GeneJet PCR Purification Kit (Fermentas, MA) and then fragmented using NEBNext dsDNA Fragmentase (New England Biolabs, MA) according to the manufacturer’s protocol, with the addition of 5 mM MgCl2 and 1 mg ml−1 BSA (New England Biolabs, MA).

    Article Title: Deletion of Snai2 and Snai3 Results in Impaired Physical Development Compounded by Lymphocyte Deficiency
    Article Snippet: Total RNA was isolated from cells using Illustra RNAspin Mini kit (GE Healthcare) according to the manufacturer’s instructions. cDNA was synthesized using SuperScript III First-Strand Synthesis System (Invitrogen) and purified with the GeneJET PCR Purification kit (Fermentas). .. Semi-quantitative RT-PCR was performed via incorporation of [32 P] dCTP [ ].

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water. .. Both of these enzymes target CCGG sequences for digestion, but only Hpa II is blocked when the inner cytosine is methylated.

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: Paragraph title: 2.5. DNA extraction, PCR and Sequencing ... PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer.

    Article Title: Geographic mosaic of symbiont selectivity in a genus of epiphytic cyanolichens
    Article Snippet: Paragraph title: DNA extraction, amplification, and sequencing ... Amplification products were purified with the GeneJET PCR Purification Kit (Fermentas).

    Article Title: Gut Microbiome Associates With Lipid-Lowering Effect of Rosuvastatin in Vivo
    Article Snippet: Paragraph title: 16S rRNA Gene Sequencing ... The 16S amplicons were purified with GeneJET PCR Purification Kit (Thermo Fisher Scientific, Waltham, MA, United States) and DNA library was constructed by using TruSeq DNA PCR-Free Library Kit (Illumina Inc., San Diego, CA, United States).

    Article Title: Prevalence of Genetic Determinants and Phenotypic Resistance to Ciprofloxacin in Campylobacter jejuni from Lithuania
    Article Snippet: Paragraph title: DNA sequencing and sequence analysis ... The PCR amplicons were purified using the GeneJet PCR purification system (Thermo Scientific, EU).

    Construct:

    Article Title: Gut Microbiome Associates With Lipid-Lowering Effect of Rosuvastatin in Vivo
    Article Snippet: After an initial denaturation step at 98°C for 1 min, PCR amplification was carried out using 30 cycles of 98°C for 10 s, 50°C for 30 s and 72°C for 30 s, and a final extension at 72°C for 5 min. PCR reactions were performed on an ABI GeneAmp 9700 PCR system (Applied Biosystems, Foster City, CA, United States). .. The 16S amplicons were purified with GeneJET PCR Purification Kit (Thermo Fisher Scientific, Waltham, MA, United States) and DNA library was constructed by using TruSeq DNA PCR-Free Library Kit (Illumina Inc., San Diego, CA, United States). .. The libraries were then sequenced by Hiseq sequencing System (Illumina Inc., San Diego, CA, United States).

    CRISPR:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: Paragraph title: CRISPR/Cas9 knockout of TRPP2. ... PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Concentration Assay:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: RNA and protein digestions were performed by addition of 1 μl of 10 μg/μl RNase A (Roche) and incubation at RT for 25 min followed by 11 μl of 10× proteinase K buffer (100 mM Tris-HCl [pH 8.0], 500 mM NaCl, 50 mM EDTA) and 1 μg proteinase K (IBI Scientific) for 1 h at 55°C. .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122). .. Libraries were submitted for 50-bp single-end Illumina sequencing as described above.

    Article Title: An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I
    Article Snippet: Briefly, the qPCR recipe was prepared with different concentrations (100, 300, 600, and 900 nM) of forward and reverse primers, for a total of 16 different concentration combinations. .. Bull trout ITSI was amplified with the assay and purified using the GeneJET PCR Purification Kit (ThermoFisher Scientific, Waltham, MA, USA).

    Sample Prep:

    Article Title: Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine
    Article Snippet: Genomic DNA sample (100 ng–1 μg) Illumina TruSeq DNA sample preparation kit (Illumina, cat. no. FC-121-2001) Ampure XP beads (Beckman Coulter, cat. no. ) Milli-Q water Sodium hydroxide, 1.0 M (NaOH; Sigma-Aldrich, cat. no. 319511) Oxidant solution (10× solution; Cambridge Epigenetix (CEGX)). .. Alternatively, in place of Steps 12–14 and 17, the oxidant and oxidation protocol can be prepared and used as described in (ref. ) Hydroxymethyl dCTP (dhmCTP, Bioline, cat. no. BIO-39046) dNTP mix, 10 mM (NEB, cat. no. N0447S) Epitect bisulfite kit (Qiagen, cat. no. 59104) P-6 saline–sodium citrate (SSC) Micro Bio-Spin columns (Bio-Rad, cat. no. 732-6200) Pfu Turbo Cx hotstart DNA polymerase (Agilent, cat. no. 600410) Agarose (Sigma-Aldrich, cat. no. A9539) dATP, 100 mM (NEB, cat. no. N0446S) dGTP, 100 mM (NEB, cat. no. N0446S) dTTP, 100 mM (NEB, cat. no. N0446S) DNA templates (Biomers; ) DNA primers (Biomers; ) DreamTaq DNA polymerase (Thermo Scientific, cat. no. EP0701) KAPA HiFi uracil + ReadyMix (KAPA Biosystems, cat. no. KK2801) GeneJet PCR purification kit (Thermo Scientific, cat. no. K0701) Taqa1 restriction endonuclease (Taq1 RE; NEB, cat. no. R0149S) Ethanol (Sigma-Aldrich, cat. no. 32221) GelRed, 3× dye (Cambridge Bioscience, cat. no. 41001) Ammonium acetate (Fisher, cat. no. A/3440/53) Acetic acid (Sigma-Aldrich, cat. no. 320099) HPLC-grade acetonitrile (Sigma-Aldrich, cat. no. 34851)

    Chromatin Immunoprecipitation:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: RNA and protein digestions were performed by addition of 1 μl of 10 μg/μl RNase A (Roche) and incubation at RT for 25 min followed by 11 μl of 10× proteinase K buffer (100 mM Tris-HCl [pH 8.0], 500 mM NaCl, 50 mM EDTA) and 1 μg proteinase K (IBI Scientific) for 1 h at 55°C. .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122). .. Libraries were submitted for 50-bp single-end Illumina sequencing as described above.

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: However, the yields of DNA recovery varied considerably among the kits. .. The Wizard® SV Gel and PCR Clean-Up System (Promega; Pr), the GeneJET PCR Purification Kit (Thermo Fisher Scientific; Th), the PureLink® PCR Purification Kit (Invitrogen; In) and the Chromatin IP DNA Purification Kit (Active Motif; Am) performed poorly with de-crosslinked chromatin. .. Consistently, these reagents recovered less than 50% of input DNA with purified DNA even when expected DNA amounts were relatively high (10–50 ng).

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: DNAs were prepared to final 1 ng, 5 ng, 10 ng and 50 ng in 100 μL ChIP elution buffer and were purified by 11 different purification reagents as suggested by the manufacturer except for the elution volume, which was fixed at 16 μL. .. Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ). .. The sample-to-beads ratio tested for Ba and Br were 1:1.25, 1:1.50, 1:1.75, and 1:2.

    Article Title: Genome-wide mapping of infection-induced SINE RNAs reveals a role in selective mRNA export
    Article Snippet: Paragraph title: ChIP-qPCR ... Following reversal of cross-links, DNA was purified using a PCR purification spin column (Fermentas) and resuspended in 50 μl of dH2 O; 1 to 2 μl of DNA was used for quantitative PCR (qPCR) with the DyNAmo ColorFlash SYBR green qPCR kit (Thermo Scientific) with appropriate primers ( ).

    Plasmid Preparation:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: We transfected mIMCD-3 cells with this plasmid using Lipofectamine 3000 (Thermo Fisher Scientific). .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Software:

    Article Title: Gut Microbiome Associates With Lipid-Lowering Effect of Rosuvastatin in Vivo
    Article Snippet: The 16S amplicons were purified with GeneJET PCR Purification Kit (Thermo Fisher Scientific, Waltham, MA, United States) and DNA library was constructed by using TruSeq DNA PCR-Free Library Kit (Illumina Inc., San Diego, CA, United States). .. The 16S amplicons were purified with GeneJET PCR Purification Kit (Thermo Fisher Scientific, Waltham, MA, United States) and DNA library was constructed by using TruSeq DNA PCR-Free Library Kit (Illumina Inc., San Diego, CA, United States).

    Real-time Polymerase Chain Reaction:

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: The Wizard® SV Gel and PCR Clean-Up System (Promega; Pr), the GeneJET PCR Purification Kit (Thermo Fisher Scientific; Th), the PureLink® PCR Purification Kit (Invitrogen; In) and the Chromatin IP DNA Purification Kit (Active Motif; Am) performed poorly with de-crosslinked chromatin. .. These reagents recovered 78.1% to 95.7% with 10–50 ng of purified DNA, 81.7% to 96.8% with 5 ng of DNA, and 68.1% to 82.9% with 1 ng of DNA except phenol/chloroform extraction with over 100%.

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: Using these optimized primer concentrations, we then performed a standard curve analysis to examine the sensitivity of the assay. .. The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific). .. A seven‐level serial dilution (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per 4 μl) was created in sterile TE, and each level was analyzed in six replicates.

    Article Title: An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I
    Article Snippet: The efficiency and sensitivity of the optimized assay was then assessed by analyzing a seven-level standard curve dilution created from purified qPCR product. .. Bull trout ITSI was amplified with the assay and purified using the GeneJET PCR Purification Kit (ThermoFisher Scientific, Waltham, MA, USA).

    Article Title: Genome-wide mapping of infection-induced SINE RNAs reveals a role in selective mRNA export
    Article Snippet: Cross-linked chromatin was immunoprecipitated using anti-POLR3A (clone ab96328; Abcam) and normal rabbit IgG (clone 2729; Cell Signaling). .. Following reversal of cross-links, DNA was purified using a PCR purification spin column (Fermentas) and resuspended in 50 μl of dH2 O; 1 to 2 μl of DNA was used for quantitative PCR (qPCR) with the DyNAmo ColorFlash SYBR green qPCR kit (Thermo Scientific) with appropriate primers ( ). .. Signals obtained by qPCR were normalized to the input DNA.

    shRNA:

    Article Title: Target discovery screens using pooled shRNA libraries and next-generation sequencing: A model workflow and analytical algorithm
    Article Snippet: Paragraph title: Genomic DNA isolation and shRNA PCR amplification ... Parallel reaction products were pooled and purified using the GeneJET PCR Purification Kit (Thermo Fisher Scientific).

    Agarose Gel Electrophoresis:

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: The other reagents MgCl2 , dNTPs, forward and reverse primers, Dream Taq Polymerase (Thermo-Scientific) and cycling conditions were optimized accordingly as shown in . .. PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer. .. Contigs were assembled using Lasergene 11 Core Suite, the DNA sequences and the predicted amino acid sequences were analysed using CLUSTAL W available in EBI website ( ) and searched by BLAST© software available at the NCBI Website and PlasmoDB version 11.0 ( ).

    Article Title: Genotyping of Plant and Animal Samples without Prior DNA Purification
    Article Snippet: Therefore, it may be necessary to either dilute ( e.g. 1:2 or 1:3 in water) or purify the PCR product before the subsequent digestion, for example by using a suitable commercial kit such as Thermo Scientific GeneJET PCR Purification Kit. .. Therefore, it may be necessary to either dilute ( e.g. 1:2 or 1:3 in water) or purify the PCR product before the subsequent digestion, for example by using a suitable commercial kit such as Thermo Scientific GeneJET PCR Purification Kit.

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: DNA templates were added to a reaction mixture containing 1.25 μl of each primer (5 pmoles), 2.5 μl buffer (Bio Basic Inc., Canada), 2 μl MgSO4 (Bio Basic Inc., Canada), 0.3 mM dNTPs, and 0.04 units/μl of taq polymerase (Bio Basic Inc., Canada). .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ). .. SNP genotyping was done by restriction fragment length polymorphism (RFLP) and confirmed by sequencing.

    Staining:

    Article Title: Nanopores Suggest a Negligible Influence of CpG Methylation on Nucleosome Packaging and Stability
    Article Snippet: Gel shift was completed by running 10 μL of the assembly reaction on a 6% polyacrylamide gel, staining the DNA with SYBR-safe (Life Technologies S33102), and imaging with a Chemi-doc imager (Biorad) ( Figure S4). .. The enzyme was then inactivated at 65 °C for 20 min. DNA was purified with the GeneJet PCR Extraction Kit (Thermo Scientific K0701) following the provided protocol except the DNA was eluted with water.

    Next-Generation Sequencing:

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ). .. The recovery rate was calculated by dividing the recovered DNA amount after purification by the starting amount and expressed in percentages.

    Knock-Out:

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: Paragraph title: CRISPR/Cas9 knockout of TRPP2. ... PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Sampling:

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific). .. The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific).

    Produced:

    Article Title: Prevalence of Genetic Determinants and Phenotypic Resistance to Ciprofloxacin in Campylobacter jejuni from Lithuania
    Article Snippet: Nested primers GZgyrA7 and GZgyrA8 (Table ), which are internal to the 673 bp gyrA PCR products produced above, were used for sequencing. .. The PCR amplicons were purified using the GeneJet PCR purification system (Thermo Scientific, EU).

    Immunoprecipitation:

    Article Title: Genome-wide mapping of infection-induced SINE RNAs reveals a role in selective mRNA export
    Article Snippet: Cross-linked chromatin was immunoprecipitated using anti-POLR3A (clone ab96328; Abcam) and normal rabbit IgG (clone 2729; Cell Signaling). .. Following reversal of cross-links, DNA was purified using a PCR purification spin column (Fermentas) and resuspended in 50 μl of dH2 O; 1 to 2 μl of DNA was used for quantitative PCR (qPCR) with the DyNAmo ColorFlash SYBR green qPCR kit (Thermo Scientific) with appropriate primers ( ).

    Magnetic Cell Separation:

    Article Title: Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells
    Article Snippet: The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122). .. The GeneJET PCR purification kit (Thermo Fisher Scientific) was used to purify the DNA, and a Qubit fluorometer in the UCR Genomics Core was used to measure the DNA concentration; 5 to 20 ng of ChIP material per condition (from one 150-mm plate) was used to generate libraries using a BIOO Scientific ChIP-seq DNA library kit (NEXTflex ChIP-seq kit catalog no. 5143-02 and barcodes catalog no. 514122).

    DNA Purification:

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: However, the yields of DNA recovery varied considerably among the kits. .. The Wizard® SV Gel and PCR Clean-Up System (Promega; Pr), the GeneJET PCR Purification Kit (Thermo Fisher Scientific; Th), the PureLink® PCR Purification Kit (Invitrogen; In) and the Chromatin IP DNA Purification Kit (Active Motif; Am) performed poorly with de-crosslinked chromatin. .. Consistently, these reagents recovered less than 50% of input DNA with purified DNA even when expected DNA amounts were relatively high (10–50 ng).

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: DNAs were prepared to final 1 ng, 5 ng, 10 ng and 50 ng in 100 μL ChIP elution buffer and were purified by 11 different purification reagents as suggested by the manufacturer except for the elution volume, which was fixed at 16 μL. .. Similarly, DNAs were purified from de-crosslinked chromatin estimated to include 1 ng, 5 ng, 10 ng, and 50 ng of DNA after treatment of RNase A and proteinase K. The following reagents were used in the experiment: ChIP DNA Clean & Concentrator™ (Cat. # D5205) from Zymo Research (Zy) (Irvine, CA); Wizard® SV Gel and PCR Clean-Up System (Cat. # A9281) from Promega (Pr) (Fitchburg, WI); GeneJET PCR Purification Kit (Cat. # K0701) from Thermo Fisher Scientific (Th) (Waltham, MA); PureLink® PCR Purification Kit (Cat. # K310001) from Invitrogen (In) (Carlsbad, CA); Monarch® PCR & DNA Cleanup Kit (Cat. # T1030S) from New England Biolabs (Ne) (Ipswich, MA); Chromatin IP DNA Purification Kit (Cat. # 58002) from Active Motif (Am) (Carlsbad, CA); QIAquick PCR Purification Kit (Cat. # 28106) from Qiagen (Qp) (Valencia, CA), MinElute PCR Purification Kit (Cat. # 28006) from Qiagen (Qm); Agencourt AMPure XP (Cat. # A63881) from Beckman (Ba) (Indianapolis, IN), RNAClean™ XP (Cat. # A63987) from Beckman (Br), and phenol/chloroform extraction (PC) (Additional file ). .. The sample-to-beads ratio tested for Ba and Br were 1:1.25, 1:1.50, 1:1.75, and 1:2.

    Article Title: The native TRPP2-dependent channel of murine renal primary cilia
    Article Snippet: After expansion, genomic DNA extracted from the clones (GeneJET Genomic DNA Purification Kit, K0721, Thermo Fisher Scientific) was used in a polymerase chain reaction (PCR, Phusion Hot Start II DNA polymerase, F549, Thermo Fisher Scientific) to amplify the targeted region. .. PCR products of restriction-site mutants were purified (GeneJet PCR Purification Kit, Thermo Fisher Scientific) and sequenced (CCHMC DNA Sequencing and Genotyping Core).

    Article Title: Geographic mosaic of symbiont selectivity in a genus of epiphytic cyanolichens
    Article Snippet: DNA was extracted from minute thallus fragments of the lichen specimen using the GeneJET Genomic DNA Purification Kit (Fermentas, Helsinki, Finland). .. Amplification products were purified with the GeneJET PCR Purification Kit (Fermentas).

    Lysis:

    Article Title: Piperaquine and Lumefantrine Resistance in Plasmodium berghei ANKA associated with Increased Expression of Ca2+/H+ antiporter and Glutathione Associated Enzymes
    Article Snippet: Briefly, packed cells were re-suspended in 5 volumes of cold (4ºC) 1× erythrocyte Lysis buffer (ammonium chloride solution) for 15-30 minutes, before spinning at 2000rpm for 8 minutes to obtain the parasite pellet. .. PCR products were analysed in 1% agarose gel, purified using GeneJet™ PCR purification kit (Thermo scientific™) and then sequenced based in BigDye v3.1 using a 3730xlsequencer.

    Marker:

    Article Title: Implication of OPRM1 A118G Polymorphism in Opioids Addicts in Pakistan: In vitro and In silico Analysis
    Article Snippet: DNA templates were added to a reaction mixture containing 1.25 μl of each primer (5 pmoles), 2.5 μl buffer (Bio Basic Inc., Canada), 2 μl MgSO4 (Bio Basic Inc., Canada), 0.3 mM dNTPs, and 0.04 units/μl of taq polymerase (Bio Basic Inc., Canada). .. The following PCR profile was used: denaturation for 5 min at 94 °C, 35 cycles for 1 min at 94 °C, 1 min at primer-specific annealing temperature (58 °C), and 2 min at 72 °C followed by final incubation at 72 °C for 4 min. Once amplified, the fragments were purified with GeneJET™ PCR Purification Kit (Fermentas, USA) and confirmed by 1% agarose gel electrophoresis using 50 bp ladder DNA molecular weight marker (Fermentas, Lithuania) (Ginosar et al. ). .. SNP genotyping was done by restriction fragment length polymorphism (RFLP) and confirmed by sequencing.

    Article Title: An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I
    Article Snippet: Bull trout ITSI was amplified with the assay and purified using the GeneJET PCR Purification Kit (ThermoFisher Scientific, Waltham, MA, USA). .. The purified product was then quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA), and serially diluted into sterile TE to create a seven-level standard curve (31 250, 6 250, 1 250, 250, 50, 10, and 2 copies per reaction).

    Gel Extraction:

    Article Title: Synthetic Peptides to Target Stringent Response-Controlled Virulence in a Pseudomonas aeruginosa Murine Cutaneous Infection Model
    Article Snippet: All ligation reactions were carried out at room temperature using Thermo Scientific T4 DNA ligase. .. DNA purifications were either performed using the GeneJET PCR purification kit (Thermo Scientific) or the GeneJET Gel extraction kit (Thermo Scientific) following the manufacturer’s instructions. .. Peptides 1018 (VRLIVAVRIWRR-NH2 ) and DJK-5 (VQWRAIRVRVIR-NH2 ) were synthesized by CPC Scientific using solid-phase 9-flurenylmethoxy carbonyl (Fmoc) chemistry and purified to > 95% purity using reverse-phase high-performance liquid chromatography (HPLC).

    Hood:

    Article Title: Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept, et al. Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept
    Article Snippet: Experiments were set up inside a hood where qPCR consumables and pipettes were irradiated with UV for 1 h before setup. .. The qPCR product was purified using GeneJET PCR Purification Kit (ThermoFisher Scientific) and quantified on a Qubit 2.0 Fluorometer (ThermoFisher Scientific).

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    Thermo Fisher genejet whole blood rna purification mini kit
    Efficiency of DNA removal by DNaseI and its fusion variants during <t>RNA</t> purification procedure. DNaseDT denotes the fusion with the (HhH) 2 domain of DNase from an extremely salt tolerant bacterium Thioalkalivibrio sp. K90mix , DNaseBS denotes the fusion with homologous domain of ComEA protein from Bacillus subtilis . The upper picture represents quantitative evaluation of undigested DNA remaining in eluates (RT-qPCR without added reverse transcriptase). The lower picture represents RT-qPCR results obtained using the same eluates. In the both pictures labels on the horizontal axis indicate the used nuclease, amount of it and the dilution ratio of the eluates before reverse transcription step and quantitation of DNA. A modified protocol of <t>GeneJET™</t> Whole Blood RNA Purification Mini Kit (Thermo Fisher Scientific, #K0761) was followed and RNA purification columns supplied by the manufacturer were used. During the experiment we have purified total blood RNA. Three arbitrary blood samples were analysed. DNA digestion was performed directly on a filter of a RNA purification column, were respective DNase enzyme was loaded.
    Genejet Whole Blood Rna Purification Mini Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 96/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/genejet whole blood rna purification mini kit/product/Thermo Fisher
    Average 96 stars, based on 14 article reviews
    Price from $9.99 to $1999.99
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    99
    Thermo Fisher pcr purification kit
    TSS-EMOTE flowchart. The TSS-EMOTE assay consists of a wet-lab library preparation (panels a to g ) and in silico analyses (panel H to N). An asterisk continually marks the original 5’-base of tri-phosphorylated RNA (thin red line). a Total RNA is purified, and digested with XRN1 5’-exonuclease, which removes the vast majority of 5’ mono-phosphorylated RNA from the sample (including 16S and 23S rRNA). b and c The XRN1 treated RNA is mixed with large excess of a synthetic RNA oligo (Rp6, shown in blue), and split into two pools. Both pools receive T4 RNA ligase, but only the “+RppH” pool is co-treated with RppH, an enzyme that converts 5’ tri-phosphorylated ends to mono-phosphorylated ends, thus allowing the ligase to use them as substrates. d and e After the ligation reaction, a semi-random primer is used to reverse-transcribe the RNA and simultaneously add a 2.0 Illumina adapter (black “B”). This results in <t>cDNA</t> with a 2.0 Illumina adaptor (for reverse reads in paired-end sequencing) at the 5’-end and if the template RNA was ligated to an Rp6 oligo, then the cDNA will also have a complementary sequence to Rp6 at the 3’-end (cRp6). f <t>PCR</t> is used to specifically amplify cDNAs that carry the 2.0 Illumina adaptor and cRp6 sequences. This step moreover adds a 1.0 Illumina adaptor (for forward reads in paired-end sequencing) and a sample-specific 4-base EMOTE barcode (blue line and “XXX”, respectively) to index the molecules (different barcodes for the -RppH and + RppH pools). The barcode of the -RppH pool will designate molecules where the XRN1 treatments has been incomplete, and this information is incorporated into the in silico analysis (see below). g The barcoded DNA from various samples (and pools) can be mixed, and loaded directly into an Illumina HiSeq machine. Millions of 50 nt sequences are obtained, each of which will span the EMOTE barcode, both known and random sections of the Rp6 oligo (see Methods ), and it will reveal the first 20 nt of the native 5’-end of the ligated RNA molecule. These 20 nt are sufficient to map the vast majority of 5’-ends to a unique position on the small genomes of the bacteria in this study. However, longer Illumina reads (and thus longer mapping sequences) can be used if the TSSs are in repeated regions or if large-genome organisms, such as humans, are being examined. h The in silico pipeline input consists of stranded RNA-seq reads for one or multiple biological replicates in FASTQ format. Each replicate includes a FASTQ for the -RppH pool and another for the + RppH pool. i The FASTQ files go through EMOTE-conv software [ 51 ] that parses the reads, aligns them to the genome, and perform the quantification. Thus, for each genomic position we obtain the number of reads whose first nucleotide align at this genomic position, and on which strand it maps. The counts are further corrected for PCR biases by looking at the unique molecular identifiers (UMIs) sequences available in the unaligned part of the EMOTE read. j Quantification counts obtained for + RppH and -RppH pools are compared through a beta-binomial model that tests whether the identified 5’ ends in the + RppH pool is significantly enriched over the identified 5’ ends in the -RppH pool at a given position. The process results in a p-value that reflects our confidence in the genomic position to be enriched in the + RppH pool of the biological replicate. k The p-values of all the biological replicates are combined into a single p-value with Fisher’s method. l and m To correct the p-values for multiple testing across all genomic positions, the false discovery rate (FDR) is evaluated and only those with a FDR ≤ 0.01 are considered to be TSSs. Note also that for the FDR is only calculated for genomic positions with at least 5 detected ligation-events in at least one of the + RppH pools (UMI ≥ 5). n The TSSs then enter an annotation process that retrieve their surrounding sequence and downstream ORFs. TSSs separated by less than 5 bp are clustered together. Finally, to draw a global picture of operon structures, an independent detection of transcription terminators is operated with the software TransTermHP [ 39 ]. o Sequence of the RNA oligo Rp6 and a typical Illumina sequencing read from a TSS-EMOTE experiment. The Recognition Sequence serves as priming site for the PCR in panel F. UMI: The randomly incorporated nucleotides in the Rp6 oligo that serves to whether Illumina reads with identical Mapping Sequences originate from separate ligation events. CS: Control Sequence. EB: EMOTE barcode to index the Illumina reads. An asterisk indicates the 5’ nucleotide of the original RNA molecule
    Pcr Purification Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 590 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pcr purification kit/product/Thermo Fisher
    Average 99 stars, based on 590 article reviews
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    pcr purification kit - by Bioz Stars, 2019-12
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    Image Search Results


    Efficiency of DNA removal by DNaseI and its fusion variants during RNA purification procedure. DNaseDT denotes the fusion with the (HhH) 2 domain of DNase from an extremely salt tolerant bacterium Thioalkalivibrio sp. K90mix , DNaseBS denotes the fusion with homologous domain of ComEA protein from Bacillus subtilis . The upper picture represents quantitative evaluation of undigested DNA remaining in eluates (RT-qPCR without added reverse transcriptase). The lower picture represents RT-qPCR results obtained using the same eluates. In the both pictures labels on the horizontal axis indicate the used nuclease, amount of it and the dilution ratio of the eluates before reverse transcription step and quantitation of DNA. A modified protocol of GeneJET™ Whole Blood RNA Purification Mini Kit (Thermo Fisher Scientific, #K0761) was followed and RNA purification columns supplied by the manufacturer were used. During the experiment we have purified total blood RNA. Three arbitrary blood samples were analysed. DNA digestion was performed directly on a filter of a RNA purification column, were respective DNase enzyme was loaded.

    Journal: PLoS ONE

    Article Title: Enhancement of DNaseI Salt Tolerance by Mimicking the Domain Structure of DNase from an Extremely Halotolerant Bacterium Thioalkalivibrio sp. K90mix

    doi: 10.1371/journal.pone.0150404

    Figure Lengend Snippet: Efficiency of DNA removal by DNaseI and its fusion variants during RNA purification procedure. DNaseDT denotes the fusion with the (HhH) 2 domain of DNase from an extremely salt tolerant bacterium Thioalkalivibrio sp. K90mix , DNaseBS denotes the fusion with homologous domain of ComEA protein from Bacillus subtilis . The upper picture represents quantitative evaluation of undigested DNA remaining in eluates (RT-qPCR without added reverse transcriptase). The lower picture represents RT-qPCR results obtained using the same eluates. In the both pictures labels on the horizontal axis indicate the used nuclease, amount of it and the dilution ratio of the eluates before reverse transcription step and quantitation of DNA. A modified protocol of GeneJET™ Whole Blood RNA Purification Mini Kit (Thermo Fisher Scientific, #K0761) was followed and RNA purification columns supplied by the manufacturer were used. During the experiment we have purified total blood RNA. Three arbitrary blood samples were analysed. DNA digestion was performed directly on a filter of a RNA purification column, were respective DNase enzyme was loaded.

    Article Snippet: RNA was purified from three arbitrary human blood samples using GeneJET™ Whole Blood RNA Purification Mini Kit (Thermo Fisher Scientific, #K0761).

    Techniques: Purification, Quantitative RT-PCR, Quantitation Assay, Modification

    TSS-EMOTE flowchart. The TSS-EMOTE assay consists of a wet-lab library preparation (panels a to g ) and in silico analyses (panel H to N). An asterisk continually marks the original 5’-base of tri-phosphorylated RNA (thin red line). a Total RNA is purified, and digested with XRN1 5’-exonuclease, which removes the vast majority of 5’ mono-phosphorylated RNA from the sample (including 16S and 23S rRNA). b and c The XRN1 treated RNA is mixed with large excess of a synthetic RNA oligo (Rp6, shown in blue), and split into two pools. Both pools receive T4 RNA ligase, but only the “+RppH” pool is co-treated with RppH, an enzyme that converts 5’ tri-phosphorylated ends to mono-phosphorylated ends, thus allowing the ligase to use them as substrates. d and e After the ligation reaction, a semi-random primer is used to reverse-transcribe the RNA and simultaneously add a 2.0 Illumina adapter (black “B”). This results in cDNA with a 2.0 Illumina adaptor (for reverse reads in paired-end sequencing) at the 5’-end and if the template RNA was ligated to an Rp6 oligo, then the cDNA will also have a complementary sequence to Rp6 at the 3’-end (cRp6). f PCR is used to specifically amplify cDNAs that carry the 2.0 Illumina adaptor and cRp6 sequences. This step moreover adds a 1.0 Illumina adaptor (for forward reads in paired-end sequencing) and a sample-specific 4-base EMOTE barcode (blue line and “XXX”, respectively) to index the molecules (different barcodes for the -RppH and + RppH pools). The barcode of the -RppH pool will designate molecules where the XRN1 treatments has been incomplete, and this information is incorporated into the in silico analysis (see below). g The barcoded DNA from various samples (and pools) can be mixed, and loaded directly into an Illumina HiSeq machine. Millions of 50 nt sequences are obtained, each of which will span the EMOTE barcode, both known and random sections of the Rp6 oligo (see Methods ), and it will reveal the first 20 nt of the native 5’-end of the ligated RNA molecule. These 20 nt are sufficient to map the vast majority of 5’-ends to a unique position on the small genomes of the bacteria in this study. However, longer Illumina reads (and thus longer mapping sequences) can be used if the TSSs are in repeated regions or if large-genome organisms, such as humans, are being examined. h The in silico pipeline input consists of stranded RNA-seq reads for one or multiple biological replicates in FASTQ format. Each replicate includes a FASTQ for the -RppH pool and another for the + RppH pool. i The FASTQ files go through EMOTE-conv software [ 51 ] that parses the reads, aligns them to the genome, and perform the quantification. Thus, for each genomic position we obtain the number of reads whose first nucleotide align at this genomic position, and on which strand it maps. The counts are further corrected for PCR biases by looking at the unique molecular identifiers (UMIs) sequences available in the unaligned part of the EMOTE read. j Quantification counts obtained for + RppH and -RppH pools are compared through a beta-binomial model that tests whether the identified 5’ ends in the + RppH pool is significantly enriched over the identified 5’ ends in the -RppH pool at a given position. The process results in a p-value that reflects our confidence in the genomic position to be enriched in the + RppH pool of the biological replicate. k The p-values of all the biological replicates are combined into a single p-value with Fisher’s method. l and m To correct the p-values for multiple testing across all genomic positions, the false discovery rate (FDR) is evaluated and only those with a FDR ≤ 0.01 are considered to be TSSs. Note also that for the FDR is only calculated for genomic positions with at least 5 detected ligation-events in at least one of the + RppH pools (UMI ≥ 5). n The TSSs then enter an annotation process that retrieve their surrounding sequence and downstream ORFs. TSSs separated by less than 5 bp are clustered together. Finally, to draw a global picture of operon structures, an independent detection of transcription terminators is operated with the software TransTermHP [ 39 ]. o Sequence of the RNA oligo Rp6 and a typical Illumina sequencing read from a TSS-EMOTE experiment. The Recognition Sequence serves as priming site for the PCR in panel F. UMI: The randomly incorporated nucleotides in the Rp6 oligo that serves to whether Illumina reads with identical Mapping Sequences originate from separate ligation events. CS: Control Sequence. EB: EMOTE barcode to index the Illumina reads. An asterisk indicates the 5’ nucleotide of the original RNA molecule

    Journal: BMC Genomics

    Article Title: TSS-EMOTE, a refined protocol for a more complete and less biased global mapping of transcription start sites in bacterial pathogens

    doi: 10.1186/s12864-016-3211-3

    Figure Lengend Snippet: TSS-EMOTE flowchart. The TSS-EMOTE assay consists of a wet-lab library preparation (panels a to g ) and in silico analyses (panel H to N). An asterisk continually marks the original 5’-base of tri-phosphorylated RNA (thin red line). a Total RNA is purified, and digested with XRN1 5’-exonuclease, which removes the vast majority of 5’ mono-phosphorylated RNA from the sample (including 16S and 23S rRNA). b and c The XRN1 treated RNA is mixed with large excess of a synthetic RNA oligo (Rp6, shown in blue), and split into two pools. Both pools receive T4 RNA ligase, but only the “+RppH” pool is co-treated with RppH, an enzyme that converts 5’ tri-phosphorylated ends to mono-phosphorylated ends, thus allowing the ligase to use them as substrates. d and e After the ligation reaction, a semi-random primer is used to reverse-transcribe the RNA and simultaneously add a 2.0 Illumina adapter (black “B”). This results in cDNA with a 2.0 Illumina adaptor (for reverse reads in paired-end sequencing) at the 5’-end and if the template RNA was ligated to an Rp6 oligo, then the cDNA will also have a complementary sequence to Rp6 at the 3’-end (cRp6). f PCR is used to specifically amplify cDNAs that carry the 2.0 Illumina adaptor and cRp6 sequences. This step moreover adds a 1.0 Illumina adaptor (for forward reads in paired-end sequencing) and a sample-specific 4-base EMOTE barcode (blue line and “XXX”, respectively) to index the molecules (different barcodes for the -RppH and + RppH pools). The barcode of the -RppH pool will designate molecules where the XRN1 treatments has been incomplete, and this information is incorporated into the in silico analysis (see below). g The barcoded DNA from various samples (and pools) can be mixed, and loaded directly into an Illumina HiSeq machine. Millions of 50 nt sequences are obtained, each of which will span the EMOTE barcode, both known and random sections of the Rp6 oligo (see Methods ), and it will reveal the first 20 nt of the native 5’-end of the ligated RNA molecule. These 20 nt are sufficient to map the vast majority of 5’-ends to a unique position on the small genomes of the bacteria in this study. However, longer Illumina reads (and thus longer mapping sequences) can be used if the TSSs are in repeated regions or if large-genome organisms, such as humans, are being examined. h The in silico pipeline input consists of stranded RNA-seq reads for one or multiple biological replicates in FASTQ format. Each replicate includes a FASTQ for the -RppH pool and another for the + RppH pool. i The FASTQ files go through EMOTE-conv software [ 51 ] that parses the reads, aligns them to the genome, and perform the quantification. Thus, for each genomic position we obtain the number of reads whose first nucleotide align at this genomic position, and on which strand it maps. The counts are further corrected for PCR biases by looking at the unique molecular identifiers (UMIs) sequences available in the unaligned part of the EMOTE read. j Quantification counts obtained for + RppH and -RppH pools are compared through a beta-binomial model that tests whether the identified 5’ ends in the + RppH pool is significantly enriched over the identified 5’ ends in the -RppH pool at a given position. The process results in a p-value that reflects our confidence in the genomic position to be enriched in the + RppH pool of the biological replicate. k The p-values of all the biological replicates are combined into a single p-value with Fisher’s method. l and m To correct the p-values for multiple testing across all genomic positions, the false discovery rate (FDR) is evaluated and only those with a FDR ≤ 0.01 are considered to be TSSs. Note also that for the FDR is only calculated for genomic positions with at least 5 detected ligation-events in at least one of the + RppH pools (UMI ≥ 5). n The TSSs then enter an annotation process that retrieve their surrounding sequence and downstream ORFs. TSSs separated by less than 5 bp are clustered together. Finally, to draw a global picture of operon structures, an independent detection of transcription terminators is operated with the software TransTermHP [ 39 ]. o Sequence of the RNA oligo Rp6 and a typical Illumina sequencing read from a TSS-EMOTE experiment. The Recognition Sequence serves as priming site for the PCR in panel F. UMI: The randomly incorporated nucleotides in the Rp6 oligo that serves to whether Illumina reads with identical Mapping Sequences originate from separate ligation events. CS: Control Sequence. EB: EMOTE barcode to index the Illumina reads. An asterisk indicates the 5’ nucleotide of the original RNA molecule

    Article Snippet: Finally, the cDNA was purified using a PCR-purification kit (GeneJET Gel Extraction Kit, Thermo Scientific, Milian, Vernier, Switzerland), and eluted in 50 μl Elution Buffer.

    Techniques: In Silico, Purification, Ligation, Sequencing, Polymerase Chain Reaction, RNA Sequencing Assay, Software

    DNA purification reagents vary in their ability to recover low amounts of DNA from de-crosslinked chromatin. a Recovered DNA amount by different DNA purification reagents from de-crosslinked chromatin. De-crosslinked chromatin estimated to include 1 ng range DNA in ChIP elution buffer was purified following the manufacturer’s instructions. The data were generated from triplicate DNA samples derived from three independent preparations. Zy, ChIP DNA Clean Concentrator™ (Zymo Research); Pr, Wizard® SV Gel and PCR Clean-Up System (Promega); Th, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, PureLink® PCR Purification Kit (Invitrogen); Ne, Monarch® PCR DNA Cleanup Kit (New England Biolabs); Am, Chromatin IP DNA Purification Kit (Active Motif); Qp, QIAquick PCR Purification Kit (Qiagen); Qm, MinElute PCR Purification Kit (Qiagen); Ba, Agencourt AMPure XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); Br, RNAClean™ XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); PC, phenol/chloroform extraction. b Interference of PCR amplification by purified eluent of purification reagents. 9 μL eluent was mixed with 1 μL 166 bp of Drosophila probe DNA (0.0001 ng), and the resulting mixture was used as the template in 20 μl of real-time PCR reaction. The Ct value for Drosophila probe DNA from TE buffer was set as 100%. The experiment was repeated 3 times using de-crosslinked chromatin estimated to include 1 ng of DNA. c Size profiles of DNA purified by different reagents. The DNAs purified from de-crosslinked chromatin estimated to include 50 ng range DNA was analyzed by AATI Fragment Analyzer. DNA size (bp) is shown

    Journal: BMC Genomics

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application

    doi: 10.1186/s12864-017-4371-5

    Figure Lengend Snippet: DNA purification reagents vary in their ability to recover low amounts of DNA from de-crosslinked chromatin. a Recovered DNA amount by different DNA purification reagents from de-crosslinked chromatin. De-crosslinked chromatin estimated to include 1 ng range DNA in ChIP elution buffer was purified following the manufacturer’s instructions. The data were generated from triplicate DNA samples derived from three independent preparations. Zy, ChIP DNA Clean Concentrator™ (Zymo Research); Pr, Wizard® SV Gel and PCR Clean-Up System (Promega); Th, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, PureLink® PCR Purification Kit (Invitrogen); Ne, Monarch® PCR DNA Cleanup Kit (New England Biolabs); Am, Chromatin IP DNA Purification Kit (Active Motif); Qp, QIAquick PCR Purification Kit (Qiagen); Qm, MinElute PCR Purification Kit (Qiagen); Ba, Agencourt AMPure XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); Br, RNAClean™ XP kit (Beckman, chromatin to beads ratio from 1:1.25 to 1:2); PC, phenol/chloroform extraction. b Interference of PCR amplification by purified eluent of purification reagents. 9 μL eluent was mixed with 1 μL 166 bp of Drosophila probe DNA (0.0001 ng), and the resulting mixture was used as the template in 20 μl of real-time PCR reaction. The Ct value for Drosophila probe DNA from TE buffer was set as 100%. The experiment was repeated 3 times using de-crosslinked chromatin estimated to include 1 ng of DNA. c Size profiles of DNA purified by different reagents. The DNAs purified from de-crosslinked chromatin estimated to include 50 ng range DNA was analyzed by AATI Fragment Analyzer. DNA size (bp) is shown

    Article Snippet: The Wizard® SV Gel and PCR Clean-Up System (Promega; Pr), the GeneJET PCR Purification Kit (Thermo Fisher Scientific; Th), the PureLink® PCR Purification Kit (Invitrogen; In) and the Chromatin IP DNA Purification Kit (Active Motif; Am) performed poorly with de-crosslinked chromatin.

    Techniques: DNA Purification, Chromatin Immunoprecipitation, Purification, Generated, Derivative Assay, Polymerase Chain Reaction, Amplification, Real-time Polymerase Chain Reaction