polymerase chain reaction pcr clean up kit Search Results


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  • 99
    Zymo Research zymoclean gel dna recovery kit
    Zymoclean Gel Dna Recovery Kit, supplied by Zymo Research, used in various techniques. Bioz Stars score: 99/100, based on 5708 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    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
    Genejet Pcr Purification Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 5651 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    MACHEREY NAGEL pcr clean up kit
    Host and viral <t>DNA</t> content of differently treated ASFV samples. Quantitative dual <t>PCR</t> was executed by Virotype ASFV PCR Kit. C t values of individual and averaged samples represented by colored circles and grey box respectively. Averages were calculated from numbers of samples indicated on X axis. Gray circles indicate minimum and maximum values. C t values higher than 40 (undetectable host DNA) are represented by 41. Whole genome amplification (WGA) was executed by REPLI-g Mini Kit.
    Pcr Clean Up Kit, supplied by MACHEREY NAGEL, used in various techniques. Bioz Stars score: 94/100, based on 6126 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher purelink 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, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, <t>PureLink®</t> 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
    Purelink Pcr Purification Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 4112 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore genelute pcr clean up 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, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, <t>PureLink®</t> 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
    Genelute Pcr Clean Up Kit, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 3243 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Qiagen ultraclean pcr clean up 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, GeneJET PCR Purification Kit (Thermo Fisher Scientific); In, <t>PureLink®</t> 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
    Ultraclean Pcr Clean Up Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 1693 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Qiagen qiaquick pcr purification kit
    TET1 and TET2 influence 5hmC on osteogenic genes. BMSC were cultured under normal growth conditions and treated with scramble siRNA or siRNA directed to TET1 (siTET1) or TET2 (siTET2) and genomic <t>DNA</t> purified and immunoprecipitated using an antibody to 5hmC. Recruitment of 5hmC to genomic regions was assessed by the hme-DIP analysis and normalised to the genomic input control. a Relative enrichment of 5hmC on RUNX2 transcription start site (TSS) was measured using <t>PCR.</t> b 5hmC on BMP2 TSS was measured as in ( a ). BMSC were cultured under normal growth and genomic DNA purified and immunoprecipitated using an antibody to 5hmC. Recruitment of 5hmc to genomic regions was assessed by the hme-DIP analysis and normalised to the genomic input control. c Cells were cultured under normal and osteogenic conditions. Relative enrichment of 5hmC on RUNX2 transcription start site (TSS), exon and intron regions. d Relative enrichment of 5hmC on BMP2 TSS, exon and intron regions, percentage input. Data represent mean S.E.M., n = 3 BMSC donors, * p ≤ 0.05, one-way ANOVA with multiple comparison analyses
    Qiaquick Pcr Purification Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 137129 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Promega pcr clean up system kit
    Illustrative representation of HSP 70 (234) amplification for Leishmania sp. before and after re-amplification of the <t>PCR</t> products. (A) PCR products of first amplification of HSP 70 (234) targeting analyzed by electrophoresis polyacrylamide gel stained with silver. Lanes: bp. molecular-weight marker (50 bp DNA ladder); 1. Infected Thrichomys laurentius from São Raimundo Nonato/PI; 2. Negative control of PCR reaction. (B) PCR products of re-amplification of the product obtained in the first PCR reaction. Lanes: bp. molecular-weight marker (50 bp DNA ladder); 1. Negative Thrichomys fosteri from Corumbá/MS (Positive only in kDNA); 2–5. Infected T. laurentius from São Raimundo Nonato/PI; 6. Infected T. fosteri from Corumbá/MS; 7. Negative control of PCR after re-amplification.
    Pcr Clean Up System Kit, supplied by Promega, used in various techniques. Bioz Stars score: 99/100, based on 1736 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa nucleospin gel
    Pipeline of microfluidic nested PCR 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.
    Nucleospin Gel, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 999 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Zymo Research dna clean and concentrator kit
    Effect of DAC concentration on <t>DNA</t> methylation and reactivation of p57 and p16 tumor suppressor genes . A), HL-60 cells were treated with the indicated concentrations of DAC for 48 h. Total DNA was isolated at 72 h and the LINE-1 element gene methylation status was analyzed by COBRA assay. B), HL-60 cells were treated with the indicated concentrations of DAC for 48 h and total RNA was isolated at 72 h. Gene expression of 18 S ribosomal RNA gene and p57 were analyzed by <t>RT-PCR.</t> The amount of DNA amplified during the exponential phase of PCR was analyzed by quantification of amplified DNA by an Agilent 2100 Bioanalyzer. The control cells are HL-60 with no drug treatment. Control vs DAC 100 ng/ml or 1,000 ng/ml p
    Dna Clean And Concentrator Kit, supplied by Zymo Research, used in various techniques. Bioz Stars score: 99/100, based on 1718 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa pcr clean up kit
    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain <t>DNA</t> (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time <t>RT-PCR</t> of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition
    Pcr Clean Up Kit, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 830 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher purelink pcr micro kit
    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain <t>DNA</t> (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time <t>RT-PCR</t> of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition
    Purelink Pcr Micro Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 988 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Zymo Research dna clean concentrator 5 kit
    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain <t>DNA</t> (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time <t>RT-PCR</t> of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition
    Dna Clean Concentrator 5 Kit, supplied by Zymo Research, used in various techniques. Bioz Stars score: 99/100, based on 1280 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher sequalprep normalization plate kit
    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain <t>DNA</t> (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time <t>RT-PCR</t> of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition
    Sequalprep Normalization Plate Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 807 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Zymo Research onestep pcr inhibitor removal kit
    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain <t>DNA</t> (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time <t>RT-PCR</t> of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition
    Onestep Pcr Inhibitor Removal Kit, supplied by Zymo Research, used in various techniques. Bioz Stars score: 99/100, based on 399 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Axygen axyprep mag pcr clean up kit
    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain <t>DNA</t> (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time <t>RT-PCR</t> of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition
    Axyprep Mag Pcr Clean Up Kit, supplied by Axygen, used in various techniques. Bioz Stars score: 89/100, based on 474 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher chargeswitch pcr clean up kit
    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain <t>DNA</t> (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time <t>RT-PCR</t> of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition
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    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: 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 ).

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

    Expression profile of 32 HERV-Ks in primary peripheral blood lymphocytes from five different donors. A 700-nt-long HERV-K env fragment was amplified from cellular RNA by RT-PCR and sequenced using the single-molecule, deep-sequencing SMRT PacBio platform.

    Journal: Journal of Virology

    Article Title: HIV-1 Interacts with Human Endogenous Retrovirus K (HML-2) Envelopes Derived from Human Primary Lymphocytes

    doi: 10.1128/JVI.00669-14

    Figure Lengend Snippet: Expression profile of 32 HERV-Ks in primary peripheral blood lymphocytes from five different donors. A 700-nt-long HERV-K env fragment was amplified from cellular RNA by RT-PCR and sequenced using the single-molecule, deep-sequencing SMRT PacBio platform.

    Article Snippet: To eliminate the interference of the reverse transcription primer on the subsequent PCR amplification reaction, HERV-K env RT products were purified using a PCR purification kit (Invitrogen) per the manufacturer's instructions.

    Techniques: Expressing, Amplification, Reverse Transcription Polymerase Chain Reaction, Sequencing

    Host and viral DNA content of differently treated ASFV samples. Quantitative dual PCR was executed by Virotype ASFV PCR Kit. C t values of individual and averaged samples represented by colored circles and grey box respectively. Averages were calculated from numbers of samples indicated on X axis. Gray circles indicate minimum and maximum values. C t values higher than 40 (undetectable host DNA) are represented by 41. Whole genome amplification (WGA) was executed by REPLI-g Mini Kit.

    Journal: Viruses

    Article Title: A Simple Method for Sample Preparation to Facilitate Efficient Whole-Genome Sequencing of African Swine Fever Virus

    doi: 10.3390/v11121129

    Figure Lengend Snippet: Host and viral DNA content of differently treated ASFV samples. Quantitative dual PCR was executed by Virotype ASFV PCR Kit. C t values of individual and averaged samples represented by colored circles and grey box respectively. Averages were calculated from numbers of samples indicated on X axis. Gray circles indicate minimum and maximum values. C t values higher than 40 (undetectable host DNA) are represented by 41. Whole genome amplification (WGA) was executed by REPLI-g Mini Kit.

    Article Snippet: Amplified DNA Clean Up REPLI-g samples were purified using the NucleoSpin Gel and PCR clean-up Kit (Macherey-Nagel Düren, Germany).

    Techniques: Polymerase Chain Reaction, Whole Genome Amplification

    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

    TET1 and TET2 influence 5hmC on osteogenic genes. BMSC were cultured under normal growth conditions and treated with scramble siRNA or siRNA directed to TET1 (siTET1) or TET2 (siTET2) and genomic DNA purified and immunoprecipitated using an antibody to 5hmC. Recruitment of 5hmC to genomic regions was assessed by the hme-DIP analysis and normalised to the genomic input control. a Relative enrichment of 5hmC on RUNX2 transcription start site (TSS) was measured using PCR. b 5hmC on BMP2 TSS was measured as in ( a ). BMSC were cultured under normal growth and genomic DNA purified and immunoprecipitated using an antibody to 5hmC. Recruitment of 5hmc to genomic regions was assessed by the hme-DIP analysis and normalised to the genomic input control. c Cells were cultured under normal and osteogenic conditions. Relative enrichment of 5hmC on RUNX2 transcription start site (TSS), exon and intron regions. d Relative enrichment of 5hmC on BMP2 TSS, exon and intron regions, percentage input. Data represent mean S.E.M., n = 3 BMSC donors, * p ≤ 0.05, one-way ANOVA with multiple comparison analyses

    Journal: Epigenetics & Chromatin

    Article Title: Specific functions of TET1 and TET2 in regulating mesenchymal cell lineage determination

    doi: 10.1186/s13072-018-0247-4

    Figure Lengend Snippet: TET1 and TET2 influence 5hmC on osteogenic genes. BMSC were cultured under normal growth conditions and treated with scramble siRNA or siRNA directed to TET1 (siTET1) or TET2 (siTET2) and genomic DNA purified and immunoprecipitated using an antibody to 5hmC. Recruitment of 5hmC to genomic regions was assessed by the hme-DIP analysis and normalised to the genomic input control. a Relative enrichment of 5hmC on RUNX2 transcription start site (TSS) was measured using PCR. b 5hmC on BMP2 TSS was measured as in ( a ). BMSC were cultured under normal growth and genomic DNA purified and immunoprecipitated using an antibody to 5hmC. Recruitment of 5hmc to genomic regions was assessed by the hme-DIP analysis and normalised to the genomic input control. c Cells were cultured under normal and osteogenic conditions. Relative enrichment of 5hmC on RUNX2 transcription start site (TSS), exon and intron regions. d Relative enrichment of 5hmC on BMP2 TSS, exon and intron regions, percentage input. Data represent mean S.E.M., n = 3 BMSC donors, * p ≤ 0.05, one-way ANOVA with multiple comparison analyses

    Article Snippet: Immunoprecipitated bead–DNA complex was treated with proteinase K for 3 h at 50 °C in elution buffer and hydroxymethylated DNA purified by using the Qiagen PCR clean-up kit (Qiaquick).

    Techniques: Cell Culture, Purification, Immunoprecipitation, Polymerase Chain Reaction

    NEDD8 enhances DNMT3b-dependent DNA methylation. A , pREP8-RTA target DNA was transfected into HEK 293T cells with or without NEDD8 or DNMT3b. Plasmid DNA was isolated by Hirt extraction 5 days after transfection and digested with the methylation-sensitive restriction enzyme HhaI, and the DNA was examined by Southern blotting. M , methylation-induced bands; U , unmethylated bands. B , analysis of the HhaI-digested DNA (from A ) by real-time PCR using primers surrounding an HhaI restriction site in the RTA promoter. The data are expressed as a ratio of the signal obtained from HhaI-digested versus undigested DNA. C , cells were transfected with siRNA control or siRNA against NAE1. After 72 h, RNA was isolated, and reverse transcription real-time PCR was performed. The ratio of NAE1 level relative to actin is presented. D , in vivo methylation assay was performed as in A , except that cells were also transfected with siRNA control or siRNA against NAE1. Error bars , S.D.

    Journal: The Journal of Biological Chemistry

    Article Title: De Novo DNA Methyltransferase DNMT3b Interacts with NEDD8-modified Proteins *

    doi: 10.1074/jbc.M110.155721

    Figure Lengend Snippet: NEDD8 enhances DNMT3b-dependent DNA methylation. A , pREP8-RTA target DNA was transfected into HEK 293T cells with or without NEDD8 or DNMT3b. Plasmid DNA was isolated by Hirt extraction 5 days after transfection and digested with the methylation-sensitive restriction enzyme HhaI, and the DNA was examined by Southern blotting. M , methylation-induced bands; U , unmethylated bands. B , analysis of the HhaI-digested DNA (from A ) by real-time PCR using primers surrounding an HhaI restriction site in the RTA promoter. The data are expressed as a ratio of the signal obtained from HhaI-digested versus undigested DNA. C , cells were transfected with siRNA control or siRNA against NAE1. After 72 h, RNA was isolated, and reverse transcription real-time PCR was performed. The ratio of NAE1 level relative to actin is presented. D , in vivo methylation assay was performed as in A , except that cells were also transfected with siRNA control or siRNA against NAE1. Error bars , S.D.

    Article Snippet: ChIP assays were performed essentially as described previously , except that after proteinase K, the DNA was purified with a PCR purification kit (Ultra Clean PCR clean-up kit, MO BIO Laboratories, Inc.).

    Techniques: DNA Methylation Assay, Transfection, Plasmid Preparation, Isolation, Methylation, Southern Blot, Real-time Polymerase Chain Reaction, In Vivo

    DNMT3b recruits CUL4A and NEDD8 to chromatin of a repressed promoter. DNMT3b recruits CUL4A and NEDD8 to chromatin. ChIP assays were performed on HEK 293T cells that were transfected with DNMT3b fused to GAL4 DNA binding domain (GAL4-DNMT3b) together with a plasmid that contains 5xGAL4 consensus binding sites (5xGAL4TKCAT). Endogenous NEDD8 ( A ) and CUL4A ( B ) were immunoprecipitated, and their association with the plasmid DNA as detected by real-time PCR relative to serum IgG ( C ) is presented. D , ChIP assay performed on HCT-116 or HCT 3b KO cell lines. The immunoprecipitated DNA was subjected to real-time PCR using primers for the endogenous PCDH10 promoter. The results are presented as -fold enrichment relative to control IgG. Error bars , S.D.

    Journal: The Journal of Biological Chemistry

    Article Title: De Novo DNA Methyltransferase DNMT3b Interacts with NEDD8-modified Proteins *

    doi: 10.1074/jbc.M110.155721

    Figure Lengend Snippet: DNMT3b recruits CUL4A and NEDD8 to chromatin of a repressed promoter. DNMT3b recruits CUL4A and NEDD8 to chromatin. ChIP assays were performed on HEK 293T cells that were transfected with DNMT3b fused to GAL4 DNA binding domain (GAL4-DNMT3b) together with a plasmid that contains 5xGAL4 consensus binding sites (5xGAL4TKCAT). Endogenous NEDD8 ( A ) and CUL4A ( B ) were immunoprecipitated, and their association with the plasmid DNA as detected by real-time PCR relative to serum IgG ( C ) is presented. D , ChIP assay performed on HCT-116 or HCT 3b KO cell lines. The immunoprecipitated DNA was subjected to real-time PCR using primers for the endogenous PCDH10 promoter. The results are presented as -fold enrichment relative to control IgG. Error bars , S.D.

    Article Snippet: ChIP assays were performed essentially as described previously , except that after proteinase K, the DNA was purified with a PCR purification kit (Ultra Clean PCR clean-up kit, MO BIO Laboratories, Inc.).

    Techniques: Chromatin Immunoprecipitation, Transfection, Binding Assay, Plasmid Preparation, Immunoprecipitation, Real-time Polymerase Chain Reaction

    Illustrative representation of HSP 70 (234) amplification for Leishmania sp. before and after re-amplification of the PCR products. (A) PCR products of first amplification of HSP 70 (234) targeting analyzed by electrophoresis polyacrylamide gel stained with silver. Lanes: bp. molecular-weight marker (50 bp DNA ladder); 1. Infected Thrichomys laurentius from São Raimundo Nonato/PI; 2. Negative control of PCR reaction. (B) PCR products of re-amplification of the product obtained in the first PCR reaction. Lanes: bp. molecular-weight marker (50 bp DNA ladder); 1. Negative Thrichomys fosteri from Corumbá/MS (Positive only in kDNA); 2–5. Infected T. laurentius from São Raimundo Nonato/PI; 6. Infected T. fosteri from Corumbá/MS; 7. Negative control of PCR after re-amplification.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Distinct Leishmania Species Infecting Wild Caviomorph Rodents (Rodentia: Hystricognathi) from Brazil

    doi: 10.1371/journal.pntd.0003389

    Figure Lengend Snippet: Illustrative representation of HSP 70 (234) amplification for Leishmania sp. before and after re-amplification of the PCR products. (A) PCR products of first amplification of HSP 70 (234) targeting analyzed by electrophoresis polyacrylamide gel stained with silver. Lanes: bp. molecular-weight marker (50 bp DNA ladder); 1. Infected Thrichomys laurentius from São Raimundo Nonato/PI; 2. Negative control of PCR reaction. (B) PCR products of re-amplification of the product obtained in the first PCR reaction. Lanes: bp. molecular-weight marker (50 bp DNA ladder); 1. Negative Thrichomys fosteri from Corumbá/MS (Positive only in kDNA); 2–5. Infected T. laurentius from São Raimundo Nonato/PI; 6. Infected T. fosteri from Corumbá/MS; 7. Negative control of PCR after re-amplification.

    Article Snippet: The PCR products obtained for HSP70 (234) targeting and the products positive only in kDNA targeting were purified using the Wizard SV Gel kit and PCR Clean-up System kit (Promega).

    Techniques: Amplification, Polymerase Chain Reaction, Electrophoresis, Staining, Molecular Weight, Marker, Infection, Negative Control, Mass Spectrometry

    Amplification, sequencing and analysis of exons 2, 5, 7, and 26 . Exons 2, 5, 7, and 26 were amplified by PCR using primers specific to these regions (a). L-100 bp ladder. PCR products were sequenced by Sangers method. All the 4 nucleotide differences were also present at genomic DNA level. Representative image of exon specific amplification (b). Alignment of exon specific sequencing results with human, mouse, and rat (predicted) sequence shows 2 amino acid residue changes (c).

    Journal: BioMed Research International

    Article Title: Developmental Testicular Expression, Cloning, and Characterization of Rat HDAC6 In Silico

    doi: 10.1155/2017/5170680

    Figure Lengend Snippet: Amplification, sequencing and analysis of exons 2, 5, 7, and 26 . Exons 2, 5, 7, and 26 were amplified by PCR using primers specific to these regions (a). L-100 bp ladder. PCR products were sequenced by Sangers method. All the 4 nucleotide differences were also present at genomic DNA level. Representative image of exon specific amplification (b). Alignment of exon specific sequencing results with human, mouse, and rat (predicted) sequence shows 2 amino acid residue changes (c).

    Article Snippet: PCR amplified DNA was cleaned using PCR clean-up kit following the kit protocol (Promega, Wisconsin, USA), sequenced, and verified using nucleotide BLAST tool.

    Techniques: Amplification, Sequencing, Polymerase Chain Reaction

    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: 10 μL) were purified using NucleoSpin Gel and the PCR Clean-up Kit (Takara Bio, Shiga, Japan).

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

    Effect of DAC concentration on DNA methylation and reactivation of p57 and p16 tumor suppressor genes . A), HL-60 cells were treated with the indicated concentrations of DAC for 48 h. Total DNA was isolated at 72 h and the LINE-1 element gene methylation status was analyzed by COBRA assay. B), HL-60 cells were treated with the indicated concentrations of DAC for 48 h and total RNA was isolated at 72 h. Gene expression of 18 S ribosomal RNA gene and p57 were analyzed by RT-PCR. The amount of DNA amplified during the exponential phase of PCR was analyzed by quantification of amplified DNA by an Agilent 2100 Bioanalyzer. The control cells are HL-60 with no drug treatment. Control vs DAC 100 ng/ml or 1,000 ng/ml p

    Journal: BMC Cancer

    Article Title: Importance of dose-schedule of 5-aza-2'-deoxycytidine for epigenetic therapy of cancer

    doi: 10.1186/1471-2407-8-128

    Figure Lengend Snippet: Effect of DAC concentration on DNA methylation and reactivation of p57 and p16 tumor suppressor genes . A), HL-60 cells were treated with the indicated concentrations of DAC for 48 h. Total DNA was isolated at 72 h and the LINE-1 element gene methylation status was analyzed by COBRA assay. B), HL-60 cells were treated with the indicated concentrations of DAC for 48 h and total RNA was isolated at 72 h. Gene expression of 18 S ribosomal RNA gene and p57 were analyzed by RT-PCR. The amount of DNA amplified during the exponential phase of PCR was analyzed by quantification of amplified DNA by an Agilent 2100 Bioanalyzer. The control cells are HL-60 with no drug treatment. Control vs DAC 100 ng/ml or 1,000 ng/ml p

    Article Snippet: The PCR products were then purified using the DNA Clean and Concentrator Kit (Zymo Research, CA, USA) and digested with Hinf I enzyme (New Englend BioLabs, MA, USA) according to manufacturer's protocols.

    Techniques: Concentration Assay, DNA Methylation Assay, Isolation, Methylation, Combined Bisulfite Restriction Analysis Assay, Expressing, Reverse Transcription Polymerase Chain Reaction, Amplification, Polymerase Chain Reaction

    EMSA analysis indicates that the predicted SLII is important for DNA binding. (A) Nucleotide sequences of the pRSP pagA promoter region. The sequence shown is located immediately upstream of the translation start codon ATG (uppercase bold). The −35 and −10 regions of a σ A ). The hypothesized SLI is presented in bold and SLII is in bold italic. Transcription start site A is indicated in uppercase bold italic. (B) Map of PAG4-1 and PAG4-2 fragments within PAG4 in relation to the hypothesized stem-loops. Numbers indicate the distance from the transcriptional start site. The figure is drawn to scale. (C) EMSA on a 2% agarose EABE gel with AtxA WT incubated with PAG4 fragment and unlabeled competitor of either PAG4-1 or PAG4-2 generated via PCR. An excess of 10×, 30×, 60×, and 100× was added in increasing amounts to the EMSAs. (D) Map of RPAGΔSLI and RPAGΔSLII fragments in relation to the hypothesized stem-loops on the labeled RPAG700 fragment. Numbers indicate the distance from the transcriptional start site. The figure is drawn to scale. (E) EMSA on a 2% agarose EABE gel with AtxA WT incubated with the RPAG700 fragment and unlabeled competitor of either RPAGΔSLI or RPAGΔSLII generated via PCR. Excesses of 10×, 30×, and 60× were added in increasing amounts to the EMSA mixtures.

    Journal: Journal of Bacteriology

    Article Title: Bacillus anthracis Virulence Regulator AtxA Binds Specifically to the pagA Promoter Region

    doi: 10.1128/JB.00569-19

    Figure Lengend Snippet: EMSA analysis indicates that the predicted SLII is important for DNA binding. (A) Nucleotide sequences of the pRSP pagA promoter region. The sequence shown is located immediately upstream of the translation start codon ATG (uppercase bold). The −35 and −10 regions of a σ A ). The hypothesized SLI is presented in bold and SLII is in bold italic. Transcription start site A is indicated in uppercase bold italic. (B) Map of PAG4-1 and PAG4-2 fragments within PAG4 in relation to the hypothesized stem-loops. Numbers indicate the distance from the transcriptional start site. The figure is drawn to scale. (C) EMSA on a 2% agarose EABE gel with AtxA WT incubated with PAG4 fragment and unlabeled competitor of either PAG4-1 or PAG4-2 generated via PCR. An excess of 10×, 30×, 60×, and 100× was added in increasing amounts to the EMSAs. (D) Map of RPAGΔSLI and RPAGΔSLII fragments in relation to the hypothesized stem-loops on the labeled RPAG700 fragment. Numbers indicate the distance from the transcriptional start site. The figure is drawn to scale. (E) EMSA on a 2% agarose EABE gel with AtxA WT incubated with the RPAG700 fragment and unlabeled competitor of either RPAGΔSLI or RPAGΔSLII generated via PCR. Excesses of 10×, 30×, and 60× were added in increasing amounts to the EMSA mixtures.

    Article Snippet: Unlabeled PCR products were purified using the DNA Clean & Concentrator kit (Zymo Research, Irvine, CA), following the manufacturer’s protocol.

    Techniques: Binding Assay, Sequencing, Incubation, Generated, Polymerase Chain Reaction, Labeling

    Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain DNA (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time RT-PCR of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition

    Journal: Nature Communications

    Article Title: Sertoli cell-only phenotype and scRNA-seq define PRAMEF12 as a factor essential for spermatogenesis in mice

    doi: 10.1038/s41467-019-13193-3

    Figure Lengend Snippet: Essential role of PRAMEF12 in spermatogonial differentiation. a Immunofluorescence of whole-mount testes from P2, P4, and P7 Pramef12 Null and Pramef12 Het mice after staining with antibodies to DDX4 (left) or KIT (middle) and merged (right). KIT-positive spermatogonia were rarely detected at P2 in either Pramef12 Null or Pramef12 Het tubules but were present at P4 and increased at P7 in Pramef12 Het tubules. Few KIT-positive spermatogonia (arrow) were observed in P7 Pramef12 Null tubules. Scale bar, 50 μm. b Immunohistochemistry of sections from P7 Pramef12 Null (top panels) and Pramef12 Het (bottom panels) testes after staining with antibodies to DDX4 (left) or KIT (middle) and merged with Hoechst 33342 to stain DNA (right). Arrowheads, KIT-positive cells in Pramef12 Null testes. Scale bar, 50 μm. c Quantitative real-time RT-PCR of Kit mRNA abundance in P7 Pramef12 Null and Pramef12 Het testes using β-actin as an internal load control and setting the abundance in Pramef12 Het testis to 1. Mean ± s.d, n = 3 biologically independent samples per condition. * P = 1.43E-05 by two-tailed Student’s t test. d Immunoblot of KIT protein in P7 Pramef12 Null and Pramef12 Het testes using α-tubulin as a load control. e Same as a , but with antibodies to SOHLH1, SOHLH2, and PLZF. The number of spermatogonia positive for SOHLH1 and SOHLH2 was significantly reduced in P7 Pramef12 Null tubules (top panels). Representative of n = 3 a , b , d , e independent biological replicates with similar results per condition

    Article Snippet: After linearization by digestion with DraI, the DNA fragment was purified with PCR Clean-up Kit (Clontech Laboratories) and transcribed using AmpliScribe T7-Flash Transcription Kit (Lucigen).

    Techniques: Immunofluorescence, Mouse Assay, Staining, Immunohistochemistry, Quantitative RT-PCR, Two Tailed Test