pcr cleanup kit  (TaKaRa)

 
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    Name:
    NucleoSpin Gel and PCR Clean Up Kit
    Description:
    Choose additional buffers sold separately for use with NucleoTrap and NucleoSpin Gel and PCR Clean Up kits for isolation of highly pure DNA fragments from agarose gels and aqueous solutions
    Catalog Number:
    740609.10
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    10 Each
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    DNA cleanup Buffers enzymes Accessories and components Nucleic acid purification
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    Structured Review

    TaKaRa pcr cleanup kit
    TGIRT ssDNA-seq workflow. The target DNA (2–50 ng in this work) is treated with alkaline phosphatase to remove 3′ phosphates and heat denatured prior to DNA-seq library construction. The resulting ssDNAs with 3′ OH termini are then used for TGIRT template-switching DNA synthesis coupled to Illumina read 2 reverse (R2R) DNA-seq adapter addition. In this novel reaction, the TGIRT-III enzyme (InGex) binds first to a synthetic 34-bp R2 RNA/R2R DNA heteroduplex in which the R2R DNA primer has a single nucleotide 3′ overhang that can direct TGIRT template switching by base pairing to the 3′ nucleotide of a target DNA strand. For minimally biased library preparation, the 3′-overhang nucleotide is an equimolar mixture of A, C, G, and T (denoted N). A 3′-blocking group (C3 spacer; 3′SpC3) is attached to the end of the R2 RNA oligonucleotide to prevent template-switching to that RNA. After the TGIRT enzyme extends the DNA primer to produce a DNA copy of the target DNA strand with an R2R adapter seamlessly linked to its 5′ end, a 5′ adenylated (App) Illumina read 1 reverse <t>(R1R)</t> adapter is added to its 3′ end by single-stranded DNA ligation using a Thermostable 5′ AppDNA/RNA ligase (New England Biolabs). In the workflow shown, an unique molecular identifier (UMI) is positioned at the 5′ end of the R1R DNA oligonucleotide. A final <t>PCR</t> step adds flow cell capture sites and barcodes for Illumina sequencing and sample multiplexing.
    Choose additional buffers sold separately for use with NucleoTrap and NucleoSpin Gel and PCR Clean Up kits for isolation of highly pure DNA fragments from agarose gels and aqueous solutions
    https://www.bioz.com/result/pcr cleanup kit/product/TaKaRa
    Average 98 stars, based on 15 article reviews
    Price from $9.99 to $1999.99
    pcr cleanup kit - by Bioz Stars, 2020-08
    98/100 stars

    Images

    1) Product Images from "Facile single-stranded DNA sequencing of human plasma DNA via thermostable group II intron reverse transcriptase template switching"

    Article Title: Facile single-stranded DNA sequencing of human plasma DNA via thermostable group II intron reverse transcriptase template switching

    Journal: Scientific Reports

    doi: 10.1038/s41598-017-09064-w

    TGIRT ssDNA-seq workflow. The target DNA (2–50 ng in this work) is treated with alkaline phosphatase to remove 3′ phosphates and heat denatured prior to DNA-seq library construction. The resulting ssDNAs with 3′ OH termini are then used for TGIRT template-switching DNA synthesis coupled to Illumina read 2 reverse (R2R) DNA-seq adapter addition. In this novel reaction, the TGIRT-III enzyme (InGex) binds first to a synthetic 34-bp R2 RNA/R2R DNA heteroduplex in which the R2R DNA primer has a single nucleotide 3′ overhang that can direct TGIRT template switching by base pairing to the 3′ nucleotide of a target DNA strand. For minimally biased library preparation, the 3′-overhang nucleotide is an equimolar mixture of A, C, G, and T (denoted N). A 3′-blocking group (C3 spacer; 3′SpC3) is attached to the end of the R2 RNA oligonucleotide to prevent template-switching to that RNA. After the TGIRT enzyme extends the DNA primer to produce a DNA copy of the target DNA strand with an R2R adapter seamlessly linked to its 5′ end, a 5′ adenylated (App) Illumina read 1 reverse (R1R) adapter is added to its 3′ end by single-stranded DNA ligation using a Thermostable 5′ AppDNA/RNA ligase (New England Biolabs). In the workflow shown, an unique molecular identifier (UMI) is positioned at the 5′ end of the R1R DNA oligonucleotide. A final PCR step adds flow cell capture sites and barcodes for Illumina sequencing and sample multiplexing.
    Figure Legend Snippet: TGIRT ssDNA-seq workflow. The target DNA (2–50 ng in this work) is treated with alkaline phosphatase to remove 3′ phosphates and heat denatured prior to DNA-seq library construction. The resulting ssDNAs with 3′ OH termini are then used for TGIRT template-switching DNA synthesis coupled to Illumina read 2 reverse (R2R) DNA-seq adapter addition. In this novel reaction, the TGIRT-III enzyme (InGex) binds first to a synthetic 34-bp R2 RNA/R2R DNA heteroduplex in which the R2R DNA primer has a single nucleotide 3′ overhang that can direct TGIRT template switching by base pairing to the 3′ nucleotide of a target DNA strand. For minimally biased library preparation, the 3′-overhang nucleotide is an equimolar mixture of A, C, G, and T (denoted N). A 3′-blocking group (C3 spacer; 3′SpC3) is attached to the end of the R2 RNA oligonucleotide to prevent template-switching to that RNA. After the TGIRT enzyme extends the DNA primer to produce a DNA copy of the target DNA strand with an R2R adapter seamlessly linked to its 5′ end, a 5′ adenylated (App) Illumina read 1 reverse (R1R) adapter is added to its 3′ end by single-stranded DNA ligation using a Thermostable 5′ AppDNA/RNA ligase (New England Biolabs). In the workflow shown, an unique molecular identifier (UMI) is positioned at the 5′ end of the R1R DNA oligonucleotide. A final PCR step adds flow cell capture sites and barcodes for Illumina sequencing and sample multiplexing.

    Techniques Used: DNA Sequencing, DNA Synthesis, Blocking Assay, DNA Ligation, Polymerase Chain Reaction, Flow Cytometry, Sequencing, Multiplexing

    Related Articles

    Amplification:

    Article Title: CRISPR/Cas9 knockout of HAS2 in rat chondrosarcoma chondrocytes demonstrates the requirement of hyaluronan for aggrecan retention
    Article Snippet: .. The PCR amplified product (myc Has2 ) was purified using the NucleoSpin Gel and PCR Clean-Up Kit (Clontech). myc Has2 was then subcloned into the pAdenoX-CMV-ZsGreen1 linearized vector using the Adeno-X Adenoviral System 3 Kit (Clontech) to form Adeno-ZsGreen1-myc Has2 . .. DNA sequences were verified for all PCR-amplified regions.

    Agarose Gel Electrophoresis:

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. The PCR was conducted under the following cycling conditions: 95°C for 3 min; 15 cycles of 95°C for 30 s, 55°C for 30 s, and 72°C for 1 min; and 72°C for 5 min. After agarose gel electrophoresis, gel lanes containing the amplicons of 200–1,000 bp were excised and purified using the NucleoSpin Gel and PCR Clean-up Kit. .. Tagged amplicons were pooled and analyzed in a MiSeq paired-end sequencing reaction with the v3 reagent kit (Illumina, Tokyo, Japan).

    Polymerase Chain Reaction:

    Article Title: The “speed gene” effect of myostatin arises in Thoroughbred horses due to a promoter proximal SINE insertion
    Article Snippet: .. The DNA was extracted from the gel using a NucleoSpin Gel and PCR Clean-Up Kit (TaKaRa-Clontech) as per manufacturers’ instructions. .. An In-Fusion HD Cloning enzyme master mix (TaKaRa-Clontech) was used along with a linearized pRACE vector (supplied with SMARTer RACE 5'/3' Kit).

    Article Title: TALEN-Mediated Modification of the Bovine Genome for Large-Scale Production of Human Serum Albumin
    Article Snippet: .. The generated PCR product was purified using the Nucleospin® Gel and PCR Clean-Up Kit (Clontech), cloned into a pCR™4-TOPO® TA vector (Invitrogen), and subsequently transformed into One Shot® Top10 Competent Cells (Invitrogen). .. Forty-eight (N = 48) selected clones were subjected to PCR analysis using primers M13F/M13R.

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. The 48 nested-PCR products were purified by means of the NucleoSpin Gel and PCR Clean-up Kit and subjected to the following sequencing library preparation. .. A sample without viral cDNA (i.e., pure water) was also subjected to the above microfluidic first-round and nested PCR in parallel as a negative control.

    Article Title: CRISPR/Cas9 knockout of HAS2 in rat chondrosarcoma chondrocytes demonstrates the requirement of hyaluronan for aggrecan retention
    Article Snippet: .. The PCR amplified product (myc Has2 ) was purified using the NucleoSpin Gel and PCR Clean-Up Kit (Clontech). myc Has2 was then subcloned into the pAdenoX-CMV-ZsGreen1 linearized vector using the Adeno-X Adenoviral System 3 Kit (Clontech) to form Adeno-ZsGreen1-myc Has2 . .. DNA sequences were verified for all PCR-amplified regions.

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. The PCR was conducted under the following cycling conditions: 95°C for 3 min; 15 cycles of 95°C for 30 s, 55°C for 30 s, and 72°C for 1 min; and 72°C for 5 min. After agarose gel electrophoresis, gel lanes containing the amplicons of 200–1,000 bp were excised and purified using the NucleoSpin Gel and PCR Clean-up Kit. .. Tagged amplicons were pooled and analyzed in a MiSeq paired-end sequencing reaction with the v3 reagent kit (Illumina, Tokyo, Japan).

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. 10 μL) were purified using NucleoSpin Gel and the PCR Clean-up Kit (Takara Bio, Shiga, Japan). .. The purified DNAs were eluted with 15 μL of the supplied elution buffer.

    Article Title: Biotinylation of Deoxyguanosine at the Abasic Site in Double-Stranded Oligodeoxynucleotides
    Article Snippet: .. After washing away unbound DNA, the biotinylated DNA was eluted and concentrated on NucleoSpin Gel and PCR Clean-up kit (Takara Bio, Japan). .. The purified DNA oligonucleotides were amplified by the DNA polymerase chain reaction with Taq polymerase and the sets of primers.

    Purification:

    Article Title: TALEN-Mediated Modification of the Bovine Genome for Large-Scale Production of Human Serum Albumin
    Article Snippet: .. The generated PCR product was purified using the Nucleospin® Gel and PCR Clean-Up Kit (Clontech), cloned into a pCR™4-TOPO® TA vector (Invitrogen), and subsequently transformed into One Shot® Top10 Competent Cells (Invitrogen). .. Forty-eight (N = 48) selected clones were subjected to PCR analysis using primers M13F/M13R.

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. The 48 nested-PCR products were purified by means of the NucleoSpin Gel and PCR Clean-up Kit and subjected to the following sequencing library preparation. .. A sample without viral cDNA (i.e., pure water) was also subjected to the above microfluidic first-round and nested PCR in parallel as a negative control.

    Article Title: CRISPR/Cas9 knockout of HAS2 in rat chondrosarcoma chondrocytes demonstrates the requirement of hyaluronan for aggrecan retention
    Article Snippet: .. The PCR amplified product (myc Has2 ) was purified using the NucleoSpin Gel and PCR Clean-Up Kit (Clontech). myc Has2 was then subcloned into the pAdenoX-CMV-ZsGreen1 linearized vector using the Adeno-X Adenoviral System 3 Kit (Clontech) to form Adeno-ZsGreen1-myc Has2 . .. DNA sequences were verified for all PCR-amplified regions.

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. The PCR was conducted under the following cycling conditions: 95°C for 3 min; 15 cycles of 95°C for 30 s, 55°C for 30 s, and 72°C for 1 min; and 72°C for 5 min. After agarose gel electrophoresis, gel lanes containing the amplicons of 200–1,000 bp were excised and purified using the NucleoSpin Gel and PCR Clean-up Kit. .. Tagged amplicons were pooled and analyzed in a MiSeq paired-end sequencing reaction with the v3 reagent kit (Illumina, Tokyo, Japan).

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. 10 μL) were purified using NucleoSpin Gel and the PCR Clean-up Kit (Takara Bio, Shiga, Japan). .. The purified DNAs were eluted with 15 μL of the supplied elution buffer.

    Nested PCR:

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. The 48 nested-PCR products were purified by means of the NucleoSpin Gel and PCR Clean-up Kit and subjected to the following sequencing library preparation. .. A sample without viral cDNA (i.e., pure water) was also subjected to the above microfluidic first-round and nested PCR in parallel as a negative control.

    Generated:

    Article Title: TALEN-Mediated Modification of the Bovine Genome for Large-Scale Production of Human Serum Albumin
    Article Snippet: .. The generated PCR product was purified using the Nucleospin® Gel and PCR Clean-Up Kit (Clontech), cloned into a pCR™4-TOPO® TA vector (Invitrogen), and subsequently transformed into One Shot® Top10 Competent Cells (Invitrogen). .. Forty-eight (N = 48) selected clones were subjected to PCR analysis using primers M13F/M13R.

    Sequencing:

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters
    Article Snippet: .. The 48 nested-PCR products were purified by means of the NucleoSpin Gel and PCR Clean-up Kit and subjected to the following sequencing library preparation. .. A sample without viral cDNA (i.e., pure water) was also subjected to the above microfluidic first-round and nested PCR in parallel as a negative control.

    Transformation Assay:

    Article Title: TALEN-Mediated Modification of the Bovine Genome for Large-Scale Production of Human Serum Albumin
    Article Snippet: .. The generated PCR product was purified using the Nucleospin® Gel and PCR Clean-Up Kit (Clontech), cloned into a pCR™4-TOPO® TA vector (Invitrogen), and subsequently transformed into One Shot® Top10 Competent Cells (Invitrogen). .. Forty-eight (N = 48) selected clones were subjected to PCR analysis using primers M13F/M13R.

    Plasmid Preparation:

    Article Title: TALEN-Mediated Modification of the Bovine Genome for Large-Scale Production of Human Serum Albumin
    Article Snippet: .. The generated PCR product was purified using the Nucleospin® Gel and PCR Clean-Up Kit (Clontech), cloned into a pCR™4-TOPO® TA vector (Invitrogen), and subsequently transformed into One Shot® Top10 Competent Cells (Invitrogen). .. Forty-eight (N = 48) selected clones were subjected to PCR analysis using primers M13F/M13R.

    Article Title: CRISPR/Cas9 knockout of HAS2 in rat chondrosarcoma chondrocytes demonstrates the requirement of hyaluronan for aggrecan retention
    Article Snippet: .. The PCR amplified product (myc Has2 ) was purified using the NucleoSpin Gel and PCR Clean-Up Kit (Clontech). myc Has2 was then subcloned into the pAdenoX-CMV-ZsGreen1 linearized vector using the Adeno-X Adenoviral System 3 Kit (Clontech) to form Adeno-ZsGreen1-myc Has2 . .. DNA sequences were verified for all PCR-amplified regions.

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  • 99
    TaKaRa pcr clean up
    Rat floxed alleles generated by zygote electroporation. a Schematic representation of CLICK: CRISPR with lssDNA inducing conditional knockout alleles. b Schematic approach to generate rat Vapb floxed alleles using the CRISPR/Cas system with lssDNA composed of the targeted exon flanked by two loxP sites (Additional file 11 : Figure S11). c <t>PCR</t> analysis of rat pups (#1–6) generated by CLICK, showing different types of mutations, indels, LD (via NHEJ), inversions, and floxed alleles (via HDR) (black arrow). The primer sets were used for PCR and sequence analysis (Additional file 2 : Figure S2). M: 100 bp <t>DNA</t> ladder marker. d Sequence analysis of 6 pups showing a variety of mutations, indels, inversion or LD indicated by red letters, and loxP insertions and flox indicated by orange letters. All pups were used for testing germline transmission (Additional file 15 : Table S1). Asterisks indicate pups used for testing the Cre-loxP system (Additional file 17 : Table S3)
    Pcr Clean Up, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 38 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pcr clean up/product/TaKaRa
    Average 99 stars, based on 38 article reviews
    Price from $9.99 to $1999.99
    pcr clean up - by Bioz Stars, 2020-08
    99/100 stars
      Buy from Supplier

    98
    TaKaRa pcr clean up kit
    Pipeline of microfluidic nested <t>PCR</t> followed by MiSeq amplicon sequencing (MFnPCR–MiSeq). Purification of PCR amplicons was carried out with the <t>NucleoSpin</t> Gel and PCR Clean-up Kit. Amplification by the first-round and nested PCR was performed on a 48.48 Access Array (AA) chip in a BioMark HD reader, while the third PCR for addition of a sample-specific tag and Illumina adapter was run in 96-well PCR plates on a conventional thermal cycler.
    Pcr Clean Up Kit, supplied by TaKaRa, used in various techniques. Bioz Stars score: 98/100, based on 92 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pcr clean up kit/product/TaKaRa
    Average 98 stars, based on 92 article reviews
    Price from $9.99 to $1999.99
    pcr clean up kit - by Bioz Stars, 2020-08
    98/100 stars
      Buy from Supplier

    Image Search Results


    Rat floxed alleles generated by zygote electroporation. a Schematic representation of CLICK: CRISPR with lssDNA inducing conditional knockout alleles. b Schematic approach to generate rat Vapb floxed alleles using the CRISPR/Cas system with lssDNA composed of the targeted exon flanked by two loxP sites (Additional file 11 : Figure S11). c PCR analysis of rat pups (#1–6) generated by CLICK, showing different types of mutations, indels, LD (via NHEJ), inversions, and floxed alleles (via HDR) (black arrow). The primer sets were used for PCR and sequence analysis (Additional file 2 : Figure S2). M: 100 bp DNA ladder marker. d Sequence analysis of 6 pups showing a variety of mutations, indels, inversion or LD indicated by red letters, and loxP insertions and flox indicated by orange letters. All pups were used for testing germline transmission (Additional file 15 : Table S1). Asterisks indicate pups used for testing the Cre-loxP system (Additional file 17 : Table S3)

    Journal: BMC Genomics

    Article Title: CLICK: one-step generation of conditional knockout mice

    doi: 10.1186/s12864-018-4713-y

    Figure Lengend Snippet: Rat floxed alleles generated by zygote electroporation. a Schematic representation of CLICK: CRISPR with lssDNA inducing conditional knockout alleles. b Schematic approach to generate rat Vapb floxed alleles using the CRISPR/Cas system with lssDNA composed of the targeted exon flanked by two loxP sites (Additional file 11 : Figure S11). c PCR analysis of rat pups (#1–6) generated by CLICK, showing different types of mutations, indels, LD (via NHEJ), inversions, and floxed alleles (via HDR) (black arrow). The primer sets were used for PCR and sequence analysis (Additional file 2 : Figure S2). M: 100 bp DNA ladder marker. d Sequence analysis of 6 pups showing a variety of mutations, indels, inversion or LD indicated by red letters, and loxP insertions and flox indicated by orange letters. All pups were used for testing germline transmission (Additional file 15 : Table S1). Asterisks indicate pups used for testing the Cre-loxP system (Additional file 17 : Table S3)

    Article Snippet: Bands corresponding to a single-strand DNA fragment were extracted using NucleoSpin® Gel and PCR Clean-up (Takara Bio, Shiga, Japan).

    Techniques: Generated, Electroporation, CRISPR, Knock-Out, Polymerase Chain Reaction, Non-Homologous End Joining, Sequencing, Marker, Transmission Assay

    One-step generation of conditional knockout animals (F0) by CLICK. a Schematic representation of applying CLICK in oocytes for in vitro fertilization (IVF) with Cre-driver mice, Emx1-cre, resulting in brain-specific recombination at the targeted floxed alleles. b PCR analysis of representative delivered mouse pups by microinjection (#1–8) or electroporation (#9–15) showing different types of mutations, indels, LD, and floxed alleles (black arrow) at the targeted Serpina3n locus. The primer sets (‘small’ in Additional file 2 : Figure S2) were used for PCR and sequence analysis. c Genotyping in several tissues: cerebrum (Cr), cerebellum (Cl), heart (H), liver (L), spleen (S), and testis (Ts), indicating recombination (red arrow) by brain-specific Cre expression in #14 mouse carrying heterozygous floxed with LD alleles. M: 100 bp DNA ladder marker. d Time-schedule comparisons of targeting methods using ES cells by CRISPR in B6 oocytes, and CLICK in Cre oocytes. CLICK saves about 6 months of crossing and reduces breeding costs during the study period (Additional file 13 : Figure S13)

    Journal: BMC Genomics

    Article Title: CLICK: one-step generation of conditional knockout mice

    doi: 10.1186/s12864-018-4713-y

    Figure Lengend Snippet: One-step generation of conditional knockout animals (F0) by CLICK. a Schematic representation of applying CLICK in oocytes for in vitro fertilization (IVF) with Cre-driver mice, Emx1-cre, resulting in brain-specific recombination at the targeted floxed alleles. b PCR analysis of representative delivered mouse pups by microinjection (#1–8) or electroporation (#9–15) showing different types of mutations, indels, LD, and floxed alleles (black arrow) at the targeted Serpina3n locus. The primer sets (‘small’ in Additional file 2 : Figure S2) were used for PCR and sequence analysis. c Genotyping in several tissues: cerebrum (Cr), cerebellum (Cl), heart (H), liver (L), spleen (S), and testis (Ts), indicating recombination (red arrow) by brain-specific Cre expression in #14 mouse carrying heterozygous floxed with LD alleles. M: 100 bp DNA ladder marker. d Time-schedule comparisons of targeting methods using ES cells by CRISPR in B6 oocytes, and CLICK in Cre oocytes. CLICK saves about 6 months of crossing and reduces breeding costs during the study period (Additional file 13 : Figure S13)

    Article Snippet: Bands corresponding to a single-strand DNA fragment were extracted using NucleoSpin® Gel and PCR Clean-up (Takara Bio, Shiga, Japan).

    Techniques: Knock-Out, In Vitro, Mouse Assay, Polymerase Chain Reaction, Electroporation, Sequencing, Expressing, Marker, CRISPR

    Mouse floxed alleles generated by microinjection of two gRNA, Cas9 mRNA and lssDNA into zygotes. a An approach to generate Serpina3n floxed alleles using the CRISPR/Cas system with long single-stranded DNA (lssDNA) composed of the targeted exon flanked by two loxP sites (Additional file 2 : Figure S2). b PCR analysis of representative delivered mouse pups (#1–15) showing different types of mutations, indels, LD, and floxed alleles (black arrow) at the targeted Serpina3n locus. The primer sets (‘small’ in Additional file 2 : Figure S2) were used for PCR and sequence analysis. Asterisks indicate pups used for testing germline transmission (Additional file 15 : Table S1). M: 100 bp DNA ladder marker. c Representative examples of the targeted Serpina3n loci generated by the microinjection of two gRNA (gRNA-1 and gRNA-2), Cas9 mRNA and lssDNA into B6 mouse zygotes (Additional file 3 : Figure S3)

    Journal: BMC Genomics

    Article Title: CLICK: one-step generation of conditional knockout mice

    doi: 10.1186/s12864-018-4713-y

    Figure Lengend Snippet: Mouse floxed alleles generated by microinjection of two gRNA, Cas9 mRNA and lssDNA into zygotes. a An approach to generate Serpina3n floxed alleles using the CRISPR/Cas system with long single-stranded DNA (lssDNA) composed of the targeted exon flanked by two loxP sites (Additional file 2 : Figure S2). b PCR analysis of representative delivered mouse pups (#1–15) showing different types of mutations, indels, LD, and floxed alleles (black arrow) at the targeted Serpina3n locus. The primer sets (‘small’ in Additional file 2 : Figure S2) were used for PCR and sequence analysis. Asterisks indicate pups used for testing germline transmission (Additional file 15 : Table S1). M: 100 bp DNA ladder marker. c Representative examples of the targeted Serpina3n loci generated by the microinjection of two gRNA (gRNA-1 and gRNA-2), Cas9 mRNA and lssDNA into B6 mouse zygotes (Additional file 3 : Figure S3)

    Article Snippet: Bands corresponding to a single-strand DNA fragment were extracted using NucleoSpin® Gel and PCR Clean-up (Takara Bio, Shiga, Japan).

    Techniques: Generated, CRISPR, Polymerase Chain Reaction, Sequencing, Transmission Assay, Marker

    Schematic of iv TRT and ssDNA preparation steps. A dsDNA template can be a PCR product, or a plasmid with a suitable restriction enzyme (RE) site distal to the insertion cassette (Steps 1–2) . The gel on the right shows a plasmid digested with a RE. RNA is synthesized using in vitro transcription (Steps 3–18) . The gel on the right shows an RNA of ~ 900 bases long. cDNA is synthesized by reverse transcription using a reverse primer (Steps 19–23) . The gel on the right shows a sample ssDNA (cDNA). Note that the cDNA preparation typically runs like a smear with a prominent band within the smear. Purification of ssDNA (Steps 24–35) . The image on the left shows the gel after excision of the prominent band (for purification) and the gel on the right shows the purified ssDNA. The GeneRuler DNA Ladder Mix (ThermoFisher Scientific, cat. no. SM0331) was used in all the gels as a DNA size marker.

    Journal: Nature protocols

    Article Title: Easi-CRISPR protocol for creating knock-in and conditional knockout mouse models using long ssDNA donors

    doi: 10.1038/nprot.2017.153

    Figure Lengend Snippet: Schematic of iv TRT and ssDNA preparation steps. A dsDNA template can be a PCR product, or a plasmid with a suitable restriction enzyme (RE) site distal to the insertion cassette (Steps 1–2) . The gel on the right shows a plasmid digested with a RE. RNA is synthesized using in vitro transcription (Steps 3–18) . The gel on the right shows an RNA of ~ 900 bases long. cDNA is synthesized by reverse transcription using a reverse primer (Steps 19–23) . The gel on the right shows a sample ssDNA (cDNA). Note that the cDNA preparation typically runs like a smear with a prominent band within the smear. Purification of ssDNA (Steps 24–35) . The image on the left shows the gel after excision of the prominent band (for purification) and the gel on the right shows the purified ssDNA. The GeneRuler DNA Ladder Mix (ThermoFisher Scientific, cat. no. SM0331) was used in all the gels as a DNA size marker.

    Article Snippet: 28) Extract DNA from the gel slice using NucleoSpin Gel or PCR Clean-up (TaKaRa) kit (option A) or by Phenol extraction and ethanol precipitation (option B).

    Techniques: Polymerase Chain Reaction, Plasmid Preparation, Synthesized, In Vitro, Purification, Marker

    Pipeline of microfluidic nested PCR followed by MiSeq amplicon sequencing (MFnPCR–MiSeq). Purification of PCR amplicons was carried out with the NucleoSpin Gel and PCR Clean-up Kit. Amplification by the first-round and nested PCR was performed on a 48.48 Access Array (AA) chip in a BioMark HD reader, while the third PCR for addition of a sample-specific tag and Illumina adapter was run in 96-well PCR plates on a conventional thermal cycler.

    Journal: Frontiers in Microbiology

    Article Title: Microfluidic PCR Amplification and MiSeq Amplicon Sequencing Techniques for High-Throughput Detection and Genotyping of Human Pathogenic RNA Viruses in Human Feces, Sewage, and Oysters

    doi: 10.3389/fmicb.2018.00830

    Figure Lengend Snippet: Pipeline of microfluidic nested PCR followed by MiSeq amplicon sequencing (MFnPCR–MiSeq). Purification of PCR amplicons was carried out with the NucleoSpin Gel and PCR Clean-up Kit. Amplification by the first-round and nested PCR was performed on a 48.48 Access Array (AA) chip in a BioMark HD reader, while the third PCR for addition of a sample-specific tag and Illumina adapter was run in 96-well PCR plates on a conventional thermal cycler.

    Article Snippet: The 48 nested-PCR products were purified by means of the NucleoSpin Gel and PCR Clean-up Kit and subjected to the following sequencing library preparation.

    Techniques: Nested PCR, Amplification, Sequencing, Purification, Polymerase Chain Reaction, Chromatin Immunoprecipitation