Facile single-stranded DNA sequencing of human plasma DNA via thermostable group II intron reverse transcriptase template switching.

ABSTRACT: High-throughput single-stranded DNA sequencing (ssDNA-seq) of cell-free DNA from plasma and other bodily fluids is a powerful method for non-invasive prenatal testing, and diagnosis of cancers and other diseases. Here, we developed a facile ssDNA-seq method, which exploits a novel template-switching activity of thermostable group II intron reverse transcriptases (TGIRTs) for DNA-seq library construction. This activity enables TGIRT enzymes to initiate DNA synthesis directly at the 3' end of a DNA strand while simultaneously attaching a DNA-seq adapter without end repair, tailing, or ligation. Initial experiments using this method to sequence E. coli genomic DNA showed that the TGIRT enzyme has surprisingly robust DNA polymerase activity. Further experiments showed that TGIRT-seq of plasma

SUBMITTER: Wu DC 

PROVIDER: S-EPMC5566474 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature