COMPOSITIONS AND METHODS FOR TARGETED NUCLEIC ACID SEQUENCE ENRICHMENT AND HIGH EFFICIENCY LIBRARY GENERATION

摘要

Provided herein are methods, compositions and kits for targeted nucleic acid sequence enrichment in a nucleic acid sample and for high efficiency nucleic acid library generation for next generation sequencing (NGS). The methods, compositions and kits provided herein can be useful for the production and capture of amplification-ready, target-specific and strand-specific regions of interest from nucleic acid samples containing complex DNA.

主权项

1. A method for enriching for a nucleic acid sequence of interest in a sample comprising nucleic acids, the method comprising:
a) annealing an oligonucleotide to the nucleic acid sequence of interest in a nucleic acid fragment, wherein the oligonucleotide comprises a 3′ portion complementary to the nucleic acid sequence of interest and a 5′ portion comprising a first adaptor sequence; b) extending the oligonucleotide annealed to the nucleic acid sequence of interest in the nucleic acid fragment with a polymerase, thereby generating an oligonucleotide extension product comprising sequence complementary to the nucleic acid sequence of interest and the first adaptor sequence at a first end; c) ligating a sequence complementary to a second adaptor sequence to the oligonucleotide extension product, thereby generating an oligonucleotide extension product comprising the first adaptor sequence at the first end, the sequence complementary to the nucleic acid sequence of interest, and a sequence complementary to the second adaptor sequence at a second end; and d) amplifying the nucleic acid sequence of interest using a first primer that anneals to the complement of the first adaptor sequence and a second primer that anneals to the complement of the second adaptor sequence to enrich for the nucleic acid sequence of interest, thereby generating an enriched nucleic acid sequence of interest.