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CRISPR-Cas9-Targeted Fragmentation and Selective Sequencing Enable Massively Parallel Microsatellite Analysis

NCJ Number
250611
Date Published
February 2017
Length
13 pages
Author(s)
GiWon Shin, Susan M. Grimes, HoJoon Lee, Billy T. Lau, Li C. Xia, Hanlee P. Ji
Agencies
NIJ-Sponsored
Annotation
This article present STR-Seq, a parallel sequencing approach that generates short tandem repeats (STRs)/microsatellite-spanning sequences, that incorporates CRISPR–Cas9 to increase the number of sequenced molecules in a targeted DNA fragment.
Abstract
Microsatellites are multi-allelic and composed of short tandem repeats (STRs) with individual motifs composed of mononucleotides, dinucleotides, or higher, including hexamers. Next-generation sequencing approaches and other STR assays rely on a limited number of PCR amplicons, typically in the tens. In their research, the authors demonstrate STR-Seq, a next-generation sequencing technology that analyses over 2,000 STRs in parallel, and provides the accurate genotyping of microsatellites. STR-Seq employs in vitro CRISPR–Cas9-targeted fragmentation to produce specific DNA molecules covering the complete microsatellite sequence. Amplification-free library preparation provides single molecule sequences without unique molecular barcodes. STR-selective primers enable massively parallel, targeted sequencing of large STR sets. Overall, STR-Seq has higher throughput, improved accuracy and provides a greater number of informative haplotypes compared with other microsatellite analysis approaches. With these new features, STR-Seq can identify a 0.1% minor genome fraction in a DNA mixture composed of different, unrelated samples. (Publisher abstract modified)
Date Created: February 16, 2017