Nested Patch PCR for highly multiplexed amplification of genomic loci.
Nested Patch polymerase chain reaction (PCR) amplifies a large number (greater than 90) of targeted loci from genomic DNA simultaneously in the same reaction. These amplified loci can then be sequenced on a second-generation sequencing machine to detect single nucleotide polymorphisms (SNPs) and mutations. The reaction is highly specific: 90% of sequencing reads match targeted loci. Nested Patch PCR can be performed on many samples in parallel, and by using sample-specific DNA barcodes, these can be pooled and sequenced in a single reaction. Thus, the Nested Patch PCR protocol that is described here provides an easy workflow to identify SNPs and mutations across many targeted loci for many samples in parallel.
Department of Genetics, Center for Genome Sciences, Washington University School of Medicine, St. Louis, MO 63108, USA.
Katherine E
KE
Varley
Robi D
RD
Mitra
D004247:DNA / Q000032:analysis*; D017931:DNA Primers / Q000235:genetics; D005821:Genetic Techniques; D023281:Genomics; D006801:Humans; D008957:Models, Genetic; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide
D017931:DNA Primers; D004247:DNA