HIGH RESOLUTION STR ANALYSIS USING NEXT GENERATION SEQUENCING

摘要

A method for analyzing short tandem repeats (STRs) is described herein. In some embodiments, the method comprises: separately digesting two portions of a genomic sample at sites that are upstream and downstream of an STR; fragmenting those products; ligating adaptors to the fragmentation products; selectively amplifying part of the top strand but not the bottom strand of the ligation products derived from the first portion, and part of the bottom strand but not the top strand of the ligation products derived from the second portion; sequencing at least some of the amplification products to produce a plurality of top strand reads and a plurality of bottom strand reads; and counting the number of STR repeats in a sequence read. A kit for performing the method is also provided.

主权项

1. A method for analyzing short tandem repeats (STRs), comprising:
(a) separately digesting, using an RNA-guided nuclease:
(i) a first portion of a genomic sample from an individual, at a defined site that is upstream of an STR; and(ii) a second portion of the sample, at a defined site that is downstream of the STR, to produce first and second digestion products; (b) fragmenting the first and second digestion products of step (a) to produce first and second fragmentation products; (c) ligating an adaptor to the fragmentation products of step (b) to produce first and second ligation products; (d) selectively amplifying, using strand-specific primers and a primer that hybridizes to the adaptor:
(i) part of the top strand but not the bottom strand of the first ligation products; and(ii) part of the bottom strand but not the top strand of the second ligation products; (e) sequencing at least some of the amplification products of step (d) to produce a plurality of top strand reads and a plurality of bottom strand reads; and (f) counting the number of STR repeats in a sequence read of step (e), thereby providing an allele-specific count of the number of STR repeats at a particular locus in the genome of the individual.