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Title: RAD cap: sequence capture of dual‐digest RAD seq libraries with identifiable duplicates and reduced missing data
Abstract

Molecular ecologists seek to genotype hundreds to thousands of loci from hundreds to thousands of individuals at minimal cost per sample. Current methods, such as restriction‐site‐associatedDNAsequencing (RADseq) and sequence capture, are constrained by costs associated with inefficient use of sequencing data and sample preparation. Here, we introduceRADcap, an approach that combines the major benefits ofRADseq (low cost with specific start positions) with those of sequence capture (repeatable sequencing of specific loci) to significantly increase efficiency and reduce costs relative to current approaches.RADcap uses a new version of dual‐digestRADseq (3RAD) to identify candidateSNPloci for capture bait design and subsequently uses custom sequence capture baits to consistently enrich candidateSNPloci across many individuals. We combined this approach with a new library preparation method for identifying and removingPCRduplicates from 3RADlibraries, which allows researchers to processRADseq data using traditional pipelines, and we tested theRADcap method by genotyping sets of 96–384Wisteriaplants. Our results demonstrate that ourRADcap method: (i) methodologically reduces (to <5%) and allows computational removal ofPCRduplicate reads from data, (ii) achieves 80–90% reads on target in 11 of 12 enrichments, (iii) returns consistent coverage (≥4×) across >90% of individuals at up to 99.8% of the targeted loci, (iv) produces consistently high occupancy matrices of genotypes across hundreds of individuals and (v) costs significantly less than current approaches.

 
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NSF-PAR ID:
10243960
Author(s) / Creator(s):
 ;  ;  ;  ;  ;  ;  
Publisher / Repository:
Wiley-Blackwell
Date Published:
Journal Name:
Molecular Ecology Resources
Volume:
16
Issue:
5
ISSN:
1755-098X
Page Range / eLocation ID:
p. 1264-1278
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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