Comparison of custom capture for targeted next-generation DNA sequencing.
Samorodnitsky E, Datta J, Jewell BM, Hagopian R, Miya J, Wing MR, Damodaran S, Lippus JM, Reeser JW, Bhatt D, Timmers CD, Roychowdhury S
J Mol Diagn 17 64-75 01/01/2015
Targeted, capture-based DNA sequencing is a cost-effective method to focus sequencing on a coding region or other customized region of the genome. There are multiple targeted sequencing methods available, but none has been systematically investigated and compared. We evaluated four commercially available custom-targeted DNA technologies for next-generation sequencing with respect to on-target sequencing, uniformity, and ability to detect single-nucleotide variations (SNVs) and copy number variations. The technologies that used sonication for DNA fragmentation displayed impressive uniformity of capture, whereas the others had shorter preparation times, but sacrificed uniformity. One of those technologies, which uses transposase for DNA fragmentation, has a drawback requiring sample pooling, and the last one, which uses restriction enzymes, has a limitation depending on restriction enzyme digest sites. Although all technologies displayed some level of concordance for calling SNVs, the technologies that require restriction enzymes or transposase missed several SNVs largely because of the lack of coverage. All technologies performed well for copy number variation calling when compared to single-nucleotide polymorphism arrays. These results enable laboratories to compare these methods to make informed decisions for their intended applications.Full Text