pcr buffer Roche Search Results


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  • 95
    Millipore tissue pcr kit
    Tissue Pcr Kit, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 25 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    82
    Roche polymerase chain reaction pcr buffer
    CD8+ cells express IL-17 within the allografts of T-bet-/- recipients. A , Intragraft expression of IL-17 was assessed by real-time <t>RT-PCR.</t> From top to bottom, <t>RNA</t> was harvested from allografts transplanted into the following mice: unmodified WT, unmodified
    Polymerase Chain Reaction Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 82/100, based on 118 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    77
    Roche 1x polymerase chain reaction pcr buffer
    CD8+ cells express IL-17 within the allografts of T-bet-/- recipients. A , Intragraft expression of IL-17 was assessed by real-time <t>RT-PCR.</t> From top to bottom, <t>RNA</t> was harvested from allografts transplanted into the following mice: unmodified WT, unmodified
    1x Polymerase Chain Reaction Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 77/100, based on 39 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    PerkinElmer polymerase chain reaction pcr buffer
    CD8+ cells express IL-17 within the allografts of T-bet-/- recipients. A , Intragraft expression of IL-17 was assessed by real-time <t>RT-PCR.</t> From top to bottom, <t>RNA</t> was harvested from allografts transplanted into the following mice: unmodified WT, unmodified
    Polymerase Chain Reaction Pcr Buffer, supplied by PerkinElmer, used in various techniques. Bioz Stars score: 85/100, based on 36 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    Roche 1×polymerase chain reaction pcr buffer
    CD8+ cells express IL-17 within the allografts of T-bet-/- recipients. A , Intragraft expression of IL-17 was assessed by real-time <t>RT-PCR.</t> From top to bottom, <t>RNA</t> was harvested from allografts transplanted into the following mice: unmodified WT, unmodified
    1×Polymerase Chain Reaction Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 79/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    Roche pcr buffer 10x roche
    CD8+ cells express IL-17 within the allografts of T-bet-/- recipients. A , Intragraft expression of IL-17 was assessed by real-time <t>RT-PCR.</t> From top to bottom, <t>RNA</t> was harvested from allografts transplanted into the following mice: unmodified WT, unmodified
    Pcr Buffer 10x Roche, supplied by Roche, used in various techniques. Bioz Stars score: 80/100, based on 53 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    Roche pcr reaction buffer roche
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Pcr Reaction Buffer Roche, supplied by Roche, used in various techniques. Bioz Stars score: 79/100, based on 33 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    84
    Roche roche faststart buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Roche Faststart Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 84/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Roche mgcl2 free buffer fast start roche
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Mgcl2 Free Buffer Fast Start Roche, supplied by Roche, used in various techniques. Bioz Stars score: 92/100, based on 21 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    81
    Roche 1x roche faststart high fidelity reaction buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    1x Roche Faststart High Fidelity Reaction Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 81/100, based on 25 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher pcr buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Pcr Buffer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1144 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    81
    Roche pcr amplification buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Pcr Amplification Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 81/100, based on 29 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    87
    Roche pcr gold buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Pcr Gold Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 87/100, based on 48 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Roche taq pcr buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Taq Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 86/100, based on 35 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Roche pcr buffer 2
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Pcr Buffer 2, supplied by Roche, used in various techniques. Bioz Stars score: 85/100, based on 44 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    82
    Roche 2x pcr buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    2x Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 82/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Roche faststart pcr buffer
    Reproducibility of <t>ISSR-PCR</t> and AP-PCR. The variation between PCR replicates, <t>DNA</t> quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.
    Faststart Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 95/100, based on 105 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    Roche roche pcr media
    Circadian oscillation profile of mPer2 in wildtype and GSK3β (-/-) ; GSK3α (flox/-) MEFs . The transcriptional profile of mPer2 and GAPDH were analyzed by reverse-transcription <t>PCR</t> in the A6(wt) and c2.1(3/4 DKO) cell lines, as depicted in panels A and B , respectively. Protein samples harvested from whole-cell lysates in parallel to RNA samples harvested for transcriptional analysis at corresponding time points were analyzed by SDS-PAGE electrophoresis. Western blot analysis of these protein samples for total-GSK3 and GAPDH is depicted in Panel C for TPs 0, 4, 12, 24, 30, and 36. Panels D and E are graphical depictions of relative levels of mPer2 expression in A6 and c2.1 based on three separately harvested A6 RNA sample sets and six c2.1 RNA sample sets. Relative values derived from densitometric measurements of PCR-amplified <t>DNA</t> bands are expressed as percentage values of mPer2 at TP1.
    Roche Pcr Media, supplied by Roche, used in various techniques. Bioz Stars score: 80/100, based on 493 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    Roche pwo pcr buffer
    Characterization of DNA analogs. ( A ) <t>PCR</t> assays analyzed by 5% native polyacrylamide gel electrophoresis. Total PCR volume 100 µl: 20 ng 418-bp DNA template (pJ1506), 0.4 mM each LJM-3222 (5'-G TA CGC AG T ) and LJM-3223 (5'-TGTGAGT AGCTCACTCAT AG ), 0.2 mM each dNTP with indicated analog triphosphate ( 1–9 ) completely replacing appropriate dNTP, and 5 U DNA polymerase (indicated with plus symbol) with associated buffer and cycle conditions. Taq DNA polymerase ( Taq ) conditions: Taq DNA polymerase buffer with 100 mg/ml BSA and 2 mM MgCl ; 98°C (3 min), 30 cycles of [94°C (30 s), 60°C (30 s), and 72°C (45 s)], 72°C (5 min). PrimeSTAR HS DNA polymerase (PS) conditions: PrimeSTAR GC buffer with 2 M betaine; 98°C (3 min), 30 cycles of [98°C (15 s), 60°C (5 s), and 72°C (45 s)], 72°C (5 min). <t>Pwo</t> SuperYield DNA Polymerase ( Pwo ) conditions: Pwo PCR buffer with GC-rich solution and 2 M betaine; 98°C (3 min), 30 cycles of [98°C (1 min), 60°C (2 min), and 72°C (8 min)], 72°C (5 min). Lane 1 is marker (M) DNA (100 bp DNA ladder, Invitrogen) with 400 - and 500-bp bands indicated. ( B ) Anion exchange chromatography of 98-bp DNA-like polymers (pJ1923). Following equilibration in 20 mM Tris–HCl, pH 8 (buffer A), samples were eluted over 25 min at a 1 ml/min flow rate in a linear gradient from 50 to 100% buffer B (buffer A plus 1 M NaCl). Eluent absorbance at 260 nm (milli-absorbance units) was monitored with elution time (min).
    Pwo Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 79/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Roche expand pcr system
    Characterization of DNA analogs. ( A ) <t>PCR</t> assays analyzed by 5% native polyacrylamide gel electrophoresis. Total PCR volume 100 µl: 20 ng 418-bp DNA template (pJ1506), 0.4 mM each LJM-3222 (5'-G TA CGC AG T ) and LJM-3223 (5'-TGTGAGT AGCTCACTCAT AG ), 0.2 mM each dNTP with indicated analog triphosphate ( 1–9 ) completely replacing appropriate dNTP, and 5 U DNA polymerase (indicated with plus symbol) with associated buffer and cycle conditions. Taq DNA polymerase ( Taq ) conditions: Taq DNA polymerase buffer with 100 mg/ml BSA and 2 mM MgCl ; 98°C (3 min), 30 cycles of [94°C (30 s), 60°C (30 s), and 72°C (45 s)], 72°C (5 min). PrimeSTAR HS DNA polymerase (PS) conditions: PrimeSTAR GC buffer with 2 M betaine; 98°C (3 min), 30 cycles of [98°C (15 s), 60°C (5 s), and 72°C (45 s)], 72°C (5 min). <t>Pwo</t> SuperYield DNA Polymerase ( Pwo ) conditions: Pwo PCR buffer with GC-rich solution and 2 M betaine; 98°C (3 min), 30 cycles of [98°C (1 min), 60°C (2 min), and 72°C (8 min)], 72°C (5 min). Lane 1 is marker (M) DNA (100 bp DNA ladder, Invitrogen) with 400 - and 500-bp bands indicated. ( B ) Anion exchange chromatography of 98-bp DNA-like polymers (pJ1923). Following equilibration in 20 mM Tris–HCl, pH 8 (buffer A), samples were eluted over 25 min at a 1 ml/min flow rate in a linear gradient from 50 to 100% buffer B (buffer A plus 1 M NaCl). Eluent absorbance at 260 nm (milli-absorbance units) was monitored with elution time (min).
    Expand Pcr System, supplied by Roche, used in various techniques. Bioz Stars score: 95/100, based on 186 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    Roche binding buffer both from high pure pcr template preparation kit roche diagnostics mannheim germany
    Characterization of DNA analogs. ( A ) <t>PCR</t> assays analyzed by 5% native polyacrylamide gel electrophoresis. Total PCR volume 100 µl: 20 ng 418-bp DNA template (pJ1506), 0.4 mM each LJM-3222 (5'-G TA CGC AG T ) and LJM-3223 (5'-TGTGAGT AGCTCACTCAT AG ), 0.2 mM each dNTP with indicated analog triphosphate ( 1–9 ) completely replacing appropriate dNTP, and 5 U DNA polymerase (indicated with plus symbol) with associated buffer and cycle conditions. Taq DNA polymerase ( Taq ) conditions: Taq DNA polymerase buffer with 100 mg/ml BSA and 2 mM MgCl ; 98°C (3 min), 30 cycles of [94°C (30 s), 60°C (30 s), and 72°C (45 s)], 72°C (5 min). PrimeSTAR HS DNA polymerase (PS) conditions: PrimeSTAR GC buffer with 2 M betaine; 98°C (3 min), 30 cycles of [98°C (15 s), 60°C (5 s), and 72°C (45 s)], 72°C (5 min). <t>Pwo</t> SuperYield DNA Polymerase ( Pwo ) conditions: Pwo PCR buffer with GC-rich solution and 2 M betaine; 98°C (3 min), 30 cycles of [98°C (1 min), 60°C (2 min), and 72°C (8 min)], 72°C (5 min). Lane 1 is marker (M) DNA (100 bp DNA ladder, Invitrogen) with 400 - and 500-bp bands indicated. ( B ) Anion exchange chromatography of 98-bp DNA-like polymers (pJ1923). Following equilibration in 20 mM Tris–HCl, pH 8 (buffer A), samples were eluted over 25 min at a 1 ml/min flow rate in a linear gradient from 50 to 100% buffer B (buffer A plus 1 M NaCl). Eluent absorbance at 260 nm (milli-absorbance units) was monitored with elution time (min).
    Binding Buffer Both From High Pure Pcr Template Preparation Kit Roche Diagnostics Mannheim Germany, supplied by Roche, used in various techniques. Bioz Stars score: 79/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Roche 1x taq pcr buffer
    Characterization of DNA analogs. ( A ) <t>PCR</t> assays analyzed by 5% native polyacrylamide gel electrophoresis. Total PCR volume 100 µl: 20 ng 418-bp DNA template (pJ1506), 0.4 mM each LJM-3222 (5'-G TA CGC AG T ) and LJM-3223 (5'-TGTGAGT AGCTCACTCAT AG ), 0.2 mM each dNTP with indicated analog triphosphate ( 1–9 ) completely replacing appropriate dNTP, and 5 U DNA polymerase (indicated with plus symbol) with associated buffer and cycle conditions. Taq DNA polymerase ( Taq ) conditions: Taq DNA polymerase buffer with 100 mg/ml BSA and 2 mM MgCl ; 98°C (3 min), 30 cycles of [94°C (30 s), 60°C (30 s), and 72°C (45 s)], 72°C (5 min). PrimeSTAR HS DNA polymerase (PS) conditions: PrimeSTAR GC buffer with 2 M betaine; 98°C (3 min), 30 cycles of [98°C (15 s), 60°C (5 s), and 72°C (45 s)], 72°C (5 min). <t>Pwo</t> SuperYield DNA Polymerase ( Pwo ) conditions: Pwo PCR buffer with GC-rich solution and 2 M betaine; 98°C (3 min), 30 cycles of [98°C (1 min), 60°C (2 min), and 72°C (8 min)], 72°C (5 min). Lane 1 is marker (M) DNA (100 bp DNA ladder, Invitrogen) with 400 - and 500-bp bands indicated. ( B ) Anion exchange chromatography of 98-bp DNA-like polymers (pJ1923). Following equilibration in 20 mM Tris–HCl, pH 8 (buffer A), samples were eluted over 25 min at a 1 ml/min flow rate in a linear gradient from 50 to 100% buffer B (buffer A plus 1 M NaCl). Eluent absorbance at 260 nm (milli-absorbance units) was monitored with elution time (min).
    1x Taq Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 88/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    82
    Roche faststart taq pcr buffer
    Characterization of DNA analogs. ( A ) <t>PCR</t> assays analyzed by 5% native polyacrylamide gel electrophoresis. Total PCR volume 100 µl: 20 ng 418-bp DNA template (pJ1506), 0.4 mM each LJM-3222 (5'-G TA CGC AG T ) and LJM-3223 (5'-TGTGAGT AGCTCACTCAT AG ), 0.2 mM each dNTP with indicated analog triphosphate ( 1–9 ) completely replacing appropriate dNTP, and 5 U DNA polymerase (indicated with plus symbol) with associated buffer and cycle conditions. Taq DNA polymerase ( Taq ) conditions: Taq DNA polymerase buffer with 100 mg/ml BSA and 2 mM MgCl ; 98°C (3 min), 30 cycles of [94°C (30 s), 60°C (30 s), and 72°C (45 s)], 72°C (5 min). PrimeSTAR HS DNA polymerase (PS) conditions: PrimeSTAR GC buffer with 2 M betaine; 98°C (3 min), 30 cycles of [98°C (15 s), 60°C (5 s), and 72°C (45 s)], 72°C (5 min). <t>Pwo</t> SuperYield DNA Polymerase ( Pwo ) conditions: Pwo PCR buffer with GC-rich solution and 2 M betaine; 98°C (3 min), 30 cycles of [98°C (1 min), 60°C (2 min), and 72°C (8 min)], 72°C (5 min). Lane 1 is marker (M) DNA (100 bp DNA ladder, Invitrogen) with 400 - and 500-bp bands indicated. ( B ) Anion exchange chromatography of 98-bp DNA-like polymers (pJ1923). Following equilibration in 20 mM Tris–HCl, pH 8 (buffer A), samples were eluted over 25 min at a 1 ml/min flow rate in a linear gradient from 50 to 100% buffer B (buffer A plus 1 M NaCl). Eluent absorbance at 260 nm (milli-absorbance units) was monitored with elution time (min).
    Faststart Taq Pcr Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 82/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Roche pcr buffer ii
    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
    Pcr Buffer Ii, supplied by Roche, used in various techniques. Bioz Stars score: 88/100, based on 143 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    75
    Roche standard pcr reaction buffer
    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
    Standard Pcr Reaction Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 75/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    Roche geneamp pcr gold buffer
    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
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    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
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    Roche pwo superyield pcr buffer
    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
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    Roche pcr reaction buffer mg2
    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
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    Roche 1x pcr reaction buffer
    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
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    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for <t>PCR</t> genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx <t>cDNA</t> indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).
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    Image Search Results


    CD8+ cells express IL-17 within the allografts of T-bet-/- recipients. A , Intragraft expression of IL-17 was assessed by real-time RT-PCR. From top to bottom, RNA was harvested from allografts transplanted into the following mice: unmodified WT, unmodified

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: CD8+ Th17 Mediate Costimulation Blockade Resistant Allograft Rejection in T-bet Deficient Mice 1

    doi:

    Figure Lengend Snippet: CD8+ cells express IL-17 within the allografts of T-bet-/- recipients. A , Intragraft expression of IL-17 was assessed by real-time RT-PCR. From top to bottom, RNA was harvested from allografts transplanted into the following mice: unmodified WT, unmodified

    Article Snippet: Five μg of total RNA were reverse transcribed using 10× PCR buffer (Roche), 10 mM dNTPs, Oligo (dT), M-MLV-RT (all from Invitrogen), and RNAsin (Promega).

    Techniques: Expressing, Quantitative RT-PCR, Mouse Assay

    Reproducibility of ISSR-PCR and AP-PCR. The variation between PCR replicates, DNA quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.

    Journal: The American Journal of Pathology

    Article Title: DNA Fingerprinting Abnormalities Can Distinguish Ulcerative Colitis Patients with Dysplasia and Cancer from Those Who Are Dysplasia/Cancer-Free

    doi:

    Figure Lengend Snippet: Reproducibility of ISSR-PCR and AP-PCR. The variation between PCR replicates, DNA quantity, DNA extraction methods, and intertissue were tested. n represents the number of samples tested, and error bars represent SD.

    Article Snippet: DNA (5 to 20 ng) was subjected to AP-PCR in 7 μl of reaction mix: 0.3 U of Taq DNA polymerase, PCR reaction buffer (Roche, Indianapolis, IN), 1.5 mmol/L MgCl2 , 200 μmol/L each dNTP, 36 ng of primers (end labeled with γ-32 P-ATP).

    Techniques: Polymerase Chain Reaction, DNA Extraction

    Cloning and characterization of a novel AP-PCR product band X. A: Fingerprint from AP-PCR. The arrow indicates the position where band X was located. B: The positions of primers and inverted Alu repeats on band X sequence. The dotted line represents the deleted sequence in high-grade dysplasia. C: Multiplex PCR of genomic DNA, the upper band indicates the presence of DNA with deletion, the lower bands indicate the presence of normal DNA. D: Multiplex PCR of microdissected DNA. NL, DNA from normal individual; F1 to F9, microdissected fractions 1 to 9.

    Journal: The American Journal of Pathology

    Article Title: DNA Fingerprinting Abnormalities Can Distinguish Ulcerative Colitis Patients with Dysplasia and Cancer from Those Who Are Dysplasia/Cancer-Free

    doi:

    Figure Lengend Snippet: Cloning and characterization of a novel AP-PCR product band X. A: Fingerprint from AP-PCR. The arrow indicates the position where band X was located. B: The positions of primers and inverted Alu repeats on band X sequence. The dotted line represents the deleted sequence in high-grade dysplasia. C: Multiplex PCR of genomic DNA, the upper band indicates the presence of DNA with deletion, the lower bands indicate the presence of normal DNA. D: Multiplex PCR of microdissected DNA. NL, DNA from normal individual; F1 to F9, microdissected fractions 1 to 9.

    Article Snippet: DNA (5 to 20 ng) was subjected to AP-PCR in 7 μl of reaction mix: 0.3 U of Taq DNA polymerase, PCR reaction buffer (Roche, Indianapolis, IN), 1.5 mmol/L MgCl2 , 200 μmol/L each dNTP, 36 ng of primers (end labeled with γ-32 P-ATP).

    Techniques: Clone Assay, Polymerase Chain Reaction, Sequencing, Multiplex Assay

    Circadian oscillation profile of mPer2 in wildtype and GSK3β (-/-) ; GSK3α (flox/-) MEFs . The transcriptional profile of mPer2 and GAPDH were analyzed by reverse-transcription PCR in the A6(wt) and c2.1(3/4 DKO) cell lines, as depicted in panels A and B , respectively. Protein samples harvested from whole-cell lysates in parallel to RNA samples harvested for transcriptional analysis at corresponding time points were analyzed by SDS-PAGE electrophoresis. Western blot analysis of these protein samples for total-GSK3 and GAPDH is depicted in Panel C for TPs 0, 4, 12, 24, 30, and 36. Panels D and E are graphical depictions of relative levels of mPer2 expression in A6 and c2.1 based on three separately harvested A6 RNA sample sets and six c2.1 RNA sample sets. Relative values derived from densitometric measurements of PCR-amplified DNA bands are expressed as percentage values of mPer2 at TP1.

    Journal: Journal of Circadian Rhythms

    Article Title: Glycogen synthase kinase 3, circadian rhythms, and bipolar disorder: a molecular link in the therapeutic action of lithium

    doi: 10.1186/1740-3391-5-3

    Figure Lengend Snippet: Circadian oscillation profile of mPer2 in wildtype and GSK3β (-/-) ; GSK3α (flox/-) MEFs . The transcriptional profile of mPer2 and GAPDH were analyzed by reverse-transcription PCR in the A6(wt) and c2.1(3/4 DKO) cell lines, as depicted in panels A and B , respectively. Protein samples harvested from whole-cell lysates in parallel to RNA samples harvested for transcriptional analysis at corresponding time points were analyzed by SDS-PAGE electrophoresis. Western blot analysis of these protein samples for total-GSK3 and GAPDH is depicted in Panel C for TPs 0, 4, 12, 24, 30, and 36. Panels D and E are graphical depictions of relative levels of mPer2 expression in A6 and c2.1 based on three separately harvested A6 RNA sample sets and six c2.1 RNA sample sets. Relative values derived from densitometric measurements of PCR-amplified DNA bands are expressed as percentage values of mPer2 at TP1.

    Article Snippet: PCR analysis of transcriptional profiles For all transcriptional profiles, PCR reactions for all time-points were prepared simultaneously, where 5 μl of cDNA was added to 45 μl of PCR mixture [10× reaction buffer, 0.5 unit Taq DNA polymerase (Roche), 0.2 mM dNTPs, and 50 pmol of primers].

    Techniques: Polymerase Chain Reaction, SDS Page, Electrophoresis, Western Blot, Expressing, Derivative Assay, Amplification

    Circadian oscillation profiles of clock genes in wild-type and GSK3β -/- MEFs . A , wild type and B , GSK3β -/- cells were synchronized, harvested, processed, and the gene products were amplified as described in the Materials and Methods. The resulting transcriptional profiles of murine GAPDH , m Per2 , m Cry1 , RevErbα , and Bmal1 were analyzed by reverse-transcription PCR. The subjective time points (TP) of peak expression are designated in white above the corresponding bands for each transcript examined. Panel C is a graphical depiction of m Per2 transcriptional oscillation based on relative values derived from densitometric measurements of PCR-amplified DNA bands in panels A and B expressed as percentages of the highest recorded value in each respective data set.

    Journal: Journal of Circadian Rhythms

    Article Title: Glycogen synthase kinase 3, circadian rhythms, and bipolar disorder: a molecular link in the therapeutic action of lithium

    doi: 10.1186/1740-3391-5-3

    Figure Lengend Snippet: Circadian oscillation profiles of clock genes in wild-type and GSK3β -/- MEFs . A , wild type and B , GSK3β -/- cells were synchronized, harvested, processed, and the gene products were amplified as described in the Materials and Methods. The resulting transcriptional profiles of murine GAPDH , m Per2 , m Cry1 , RevErbα , and Bmal1 were analyzed by reverse-transcription PCR. The subjective time points (TP) of peak expression are designated in white above the corresponding bands for each transcript examined. Panel C is a graphical depiction of m Per2 transcriptional oscillation based on relative values derived from densitometric measurements of PCR-amplified DNA bands in panels A and B expressed as percentages of the highest recorded value in each respective data set.

    Article Snippet: PCR analysis of transcriptional profiles For all transcriptional profiles, PCR reactions for all time-points were prepared simultaneously, where 5 μl of cDNA was added to 45 μl of PCR mixture [10× reaction buffer, 0.5 unit Taq DNA polymerase (Roche), 0.2 mM dNTPs, and 50 pmol of primers].

    Techniques: Amplification, Polymerase Chain Reaction, Expressing, Derivative Assay

    Circadian oscillation profiles of mPer2 following pharmacological or genetic GSK3 inhibition . The transcriptional profile mPer2 ( A ) was analyzed by reverse-transcription PCR in: wild-type, GSK3β -/- / GSK3α RNAi (clones A1.4 and A1.6); as well as 20 mM Lithium or 25 μM kenpaullone treatment in a wild-type background. The subjective time points of peak expression are designated in white above the corresponding bands for each transcript examined. The time intervals where these effects are most visible (TP0-4, TP24-32, and TP44-52) are isolated in white boxes. The effects of genetic ( B ) and pharmacological ( C ) interference of GSK3 activity on mPer2 transcriptional oscillation are graphically depicted based on relative values derived from densitometric measurements of PCR-amplified DNA bands in panel A expressed as percentages of the highest recorded value in each respective data set.

    Journal: Journal of Circadian Rhythms

    Article Title: Glycogen synthase kinase 3, circadian rhythms, and bipolar disorder: a molecular link in the therapeutic action of lithium

    doi: 10.1186/1740-3391-5-3

    Figure Lengend Snippet: Circadian oscillation profiles of mPer2 following pharmacological or genetic GSK3 inhibition . The transcriptional profile mPer2 ( A ) was analyzed by reverse-transcription PCR in: wild-type, GSK3β -/- / GSK3α RNAi (clones A1.4 and A1.6); as well as 20 mM Lithium or 25 μM kenpaullone treatment in a wild-type background. The subjective time points of peak expression are designated in white above the corresponding bands for each transcript examined. The time intervals where these effects are most visible (TP0-4, TP24-32, and TP44-52) are isolated in white boxes. The effects of genetic ( B ) and pharmacological ( C ) interference of GSK3 activity on mPer2 transcriptional oscillation are graphically depicted based on relative values derived from densitometric measurements of PCR-amplified DNA bands in panel A expressed as percentages of the highest recorded value in each respective data set.

    Article Snippet: PCR analysis of transcriptional profiles For all transcriptional profiles, PCR reactions for all time-points were prepared simultaneously, where 5 μl of cDNA was added to 45 μl of PCR mixture [10× reaction buffer, 0.5 unit Taq DNA polymerase (Roche), 0.2 mM dNTPs, and 50 pmol of primers].

    Techniques: Inhibition, Polymerase Chain Reaction, Expressing, Isolation, Activity Assay, Derivative Assay, Amplification

    Characterization of DNA analogs. ( A ) PCR assays analyzed by 5% native polyacrylamide gel electrophoresis. Total PCR volume 100 µl: 20 ng 418-bp DNA template (pJ1506), 0.4 mM each LJM-3222 (5'-G TA CGC AG T ) and LJM-3223 (5'-TGTGAGT AGCTCACTCAT AG ), 0.2 mM each dNTP with indicated analog triphosphate ( 1–9 ) completely replacing appropriate dNTP, and 5 U DNA polymerase (indicated with plus symbol) with associated buffer and cycle conditions. Taq DNA polymerase ( Taq ) conditions: Taq DNA polymerase buffer with 100 mg/ml BSA and 2 mM MgCl ; 98°C (3 min), 30 cycles of [94°C (30 s), 60°C (30 s), and 72°C (45 s)], 72°C (5 min). PrimeSTAR HS DNA polymerase (PS) conditions: PrimeSTAR GC buffer with 2 M betaine; 98°C (3 min), 30 cycles of [98°C (15 s), 60°C (5 s), and 72°C (45 s)], 72°C (5 min). Pwo SuperYield DNA Polymerase ( Pwo ) conditions: Pwo PCR buffer with GC-rich solution and 2 M betaine; 98°C (3 min), 30 cycles of [98°C (1 min), 60°C (2 min), and 72°C (8 min)], 72°C (5 min). Lane 1 is marker (M) DNA (100 bp DNA ladder, Invitrogen) with 400 - and 500-bp bands indicated. ( B ) Anion exchange chromatography of 98-bp DNA-like polymers (pJ1923). Following equilibration in 20 mM Tris–HCl, pH 8 (buffer A), samples were eluted over 25 min at a 1 ml/min flow rate in a linear gradient from 50 to 100% buffer B (buffer A plus 1 M NaCl). Eluent absorbance at 260 nm (milli-absorbance units) was monitored with elution time (min).

    Journal: Nucleic Acids Research

    Article Title: Mechanical properties of DNA-like polymers

    doi: 10.1093/nar/gkt808

    Figure Lengend Snippet: Characterization of DNA analogs. ( A ) PCR assays analyzed by 5% native polyacrylamide gel electrophoresis. Total PCR volume 100 µl: 20 ng 418-bp DNA template (pJ1506), 0.4 mM each LJM-3222 (5'-G TA CGC AG T ) and LJM-3223 (5'-TGTGAGT AGCTCACTCAT AG ), 0.2 mM each dNTP with indicated analog triphosphate ( 1–9 ) completely replacing appropriate dNTP, and 5 U DNA polymerase (indicated with plus symbol) with associated buffer and cycle conditions. Taq DNA polymerase ( Taq ) conditions: Taq DNA polymerase buffer with 100 mg/ml BSA and 2 mM MgCl ; 98°C (3 min), 30 cycles of [94°C (30 s), 60°C (30 s), and 72°C (45 s)], 72°C (5 min). PrimeSTAR HS DNA polymerase (PS) conditions: PrimeSTAR GC buffer with 2 M betaine; 98°C (3 min), 30 cycles of [98°C (15 s), 60°C (5 s), and 72°C (45 s)], 72°C (5 min). Pwo SuperYield DNA Polymerase ( Pwo ) conditions: Pwo PCR buffer with GC-rich solution and 2 M betaine; 98°C (3 min), 30 cycles of [98°C (1 min), 60°C (2 min), and 72°C (8 min)], 72°C (5 min). Lane 1 is marker (M) DNA (100 bp DNA ladder, Invitrogen) with 400 - and 500-bp bands indicated. ( B ) Anion exchange chromatography of 98-bp DNA-like polymers (pJ1923). Following equilibration in 20 mM Tris–HCl, pH 8 (buffer A), samples were eluted over 25 min at a 1 ml/min flow rate in a linear gradient from 50 to 100% buffer B (buffer A plus 1 M NaCl). Eluent absorbance at 260 nm (milli-absorbance units) was monitored with elution time (min).

    Article Snippet: For analogs 7 and 8 , PCR reactions (100 µl) included 20 ng template, 0.4 mM forward and reverse primers, Pwo PCR buffer (Roche), GC-rich solution (Roche), 0.2 mM each dNTP (dTTP completely replaced with analog), 2 M betaine (Sigma-Aldrich) and 5 U Pwo SuperYield DNA Polymerase (Roche).

    Techniques: Polymerase Chain Reaction, Polyacrylamide Gel Electrophoresis, Marker, Chromatography, Flow Cytometry

    Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for PCR genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx cDNA indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).

    Journal: PLoS Genetics

    Article Title: Senataxin Plays an Essential Role with DNA Damage Response Proteins in Meiotic Recombination and Gene Silencing

    doi: 10.1371/journal.pgen.1003435

    Figure Lengend Snippet: Targeted disruption of the mouse Setx gene. A. Diagram of the Setx wild type allele (WT), targeting vector, and mutant alleles (neo+ and KO). Primers used for PCR genotyping (In3F, In3R, LoxPR) and the length of the PCR fragments obtained for WT (In3F and In3R yielding a 600 bp product) and KO (In3F and LoxpR yielding a 339 bp product) are indicated. E, exon; I, intron. NeoR represents the neomycin cassette, and triangles the loxP sites. B. Representative image of PCR genotyping using In3F, In3R and LoxPR primers. Wild type (+/+), heterozygotes (+/−) and knockout (−/−) alleles generate PCR products of 600 bp, 600 bp and 339 bp, and 339 bp, respectively. A negative control for the PCR reaction (−ve) is also shown. M, 100 bp marker. C. RT-PCR of 35 day-old mice testes samples using primers specific to Setx cDNA indicates the absence of Setx expression in KO testes. GAPDH was used as an internal standard. D. Immunoprecipitation of senataxin using anti-human senataxin antibodies (Ab-1/Ab-3) from 35 day-old mouse testes extracts confirmed the absence of the protein in the Setx −/− . Immunoprecipitation of senataxin from human lymphoblastoid cell extracts from normal (C3ABR) and an AOA2 patient (SETX2RM) confirmed the similar size of senataxin in both species. A species-matched non-specific serum (NSIg) was used as a negative control in for the IP experiments. As expected, no senataxin protein was brought down from Setx +/+ testes following the IP with the non-specific serum (NSIg).

    Article Snippet: Reactions (25 µl) contained 14.5 µl of sterile water, 50 ng of cDNA template, 1× PCR Buffer II (Roche, Switzerland), 2.5 mM MgCl2 (Roche, Switzerland), 20 µM dNTPs, 1 µM of each primer, and 5 µl of AmpliTaq Gold DNA Polymerase (Roche, Switzerland).

    Techniques: Plasmid Preparation, Mutagenesis, Polymerase Chain Reaction, Knock-Out, Negative Control, Marker, Reverse Transcription Polymerase Chain Reaction, Mouse Assay, Expressing, Immunoprecipitation

    Differential gene expression profiles of hCSSC and keratocytes. mRNA abundance relative to 18s ribosomal RNA was determined by qRT-PCR for a panel of 18 genes using cDNA from primary human keratocytes cultured in protein-free DME-F12 medium and from hCSSC

    Journal:

    Article Title: Secretion and Organization of a Cornea-like Tissue In Vitro by Stem Cells from Human Corneal Stroma

    doi: 10.1167/iovs.07-0587

    Figure Lengend Snippet: Differential gene expression profiles of hCSSC and keratocytes. mRNA abundance relative to 18s ribosomal RNA was determined by qRT-PCR for a panel of 18 genes using cDNA from primary human keratocytes cultured in protein-free DME-F12 medium and from hCSSC

    Article Snippet: RNA (400 ng) was transcribed to cDNA in a 100- μ L reaction containing 1× PCR II buffer (Roche Applied Science, Indianapolis, IN), 5 mM MgCl2 , 200 μ M dNTP mixture (Roche), 2.5 μ M random hexamers (Invitrogen), 0.4 U RNase inhibitor, and 125 U SuperScript II reverse transcriptase (Invitrogen).

    Techniques: Expressing, Quantitative RT-PCR, Cell Culture