Theoretical and experimental assessment of degenerate primer tagging in ultra-deep applications of next-generation sequencing.

ABSTRACT: Primer IDs (pIDs) are random oligonucleotide tags used in next-generation sequencing to identify sequences that originate from the same template. These tags are produced by degenerate primers during the reverse transcription of RNA molecules into cDNA. The use of pIDs helps to track the number of RNA molecules carried through amplification and sequencing, and allows resolution of inconsistencies between reads sharing a pID. Three potential issues complicate the above applications. First, multiple cDNAs may share a pID by chance; we found that while preventing any cDNAs from sharing a pID may be unfeasible, it is still practical to limit the number of these collisions. Secondly, a pID must be observed in at least three sequences to allow error correction; as such, pIDs observed only one or

SUBMITTER: Liang RH 

PROVIDER: S-EPMC4081055 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature