Long-adapter single-strand oligonucleotide probes for the massively multiplexed cloning of kilobase genome regions.

ABSTRACT: As the catalogue of sequenced genomes and metagenomes continues to grow, massively parallel approaches for the comprehensive and functional analysis of gene products and regulatory elements are becoming increasingly valuable. Current strategies to synthesize or clone complex libraries of DNA sequences are limited by the length of the DNA targets, throughput and cost. Here, we show that long-adapter single-strand oligonucleotide (LASSO) probes can capture and clone thousands of kilobase DNA fragments in a single reaction. As a proof-of-principle, we simultaneously cloned >3,000 bacterial open reading frames (ORFs) from E. coli genomic DNA (spanning 400-5,000 bp targets). Targets were enriched up to a median of ~60-fold compared to non-targeted genomic regions. At a cutoff of 3 times

SUBMITTER: Tosi L 

PROVIDER: S-EPMC5687285 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature