Universal PCR genotyping assay that achieves single copy sensitivity with any primer pair.
JL Stratman, WM Barnes, TC Simon - 2003 - cabidigitallibrary.org
JL Stratman, WM Barnes, TC Simon
2003•cabidigitallibrary.orgConditions typically utilized for long and accurate polymerase chain reaction (PCR) were
modified to develop a sensitive and robust assay for genotyping genetically altered mice.
Primers used had lengths of 27-30 nucleotides, with 50-60% guanosine/cytosine content
and produce a 100-500 nucleotide amplicon. Genomic DNA isolated by HotSHOT was used
as template (although DNA isolated by other methods also work). Cycling conditions were
93° C (for 1 minute), followed by 30 cycles of 93° C (for 20 seconds) and 68° C (for 3 …
modified to develop a sensitive and robust assay for genotyping genetically altered mice.
Primers used had lengths of 27-30 nucleotides, with 50-60% guanosine/cytosine content
and produce a 100-500 nucleotide amplicon. Genomic DNA isolated by HotSHOT was used
as template (although DNA isolated by other methods also work). Cycling conditions were
93° C (for 1 minute), followed by 30 cycles of 93° C (for 20 seconds) and 68° C (for 3 …
Abstract
Conditions typically utilized for long and accurate polymerase chain reaction (PCR) were modified to develop a sensitive and robust assay for genotyping genetically altered mice. Primers used had lengths of 27-30 nucleotides, with 50-60% guanosine/cytosine content and produce a 100-500 nucleotide amplicon. Genomic DNA isolated by HotSHOT was used as template (although DNA isolated by other methods also work). Cycling conditions were 93°C (for 1 minute), followed by 30 cycles of 93°C (for 20 seconds) and 68°C (for 3 minutes). The simplified assay provides a single set of conditions for all primer sets, and all primers chosen work at single copy sensitivity.
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