Methods of making di-tagged DNA libraries from DNA or RNA using double-tagged oligonucleotides

摘要

Disclosed are methods, compositions and kits related to making double-tagged DNA libraries from RNA/DNA samples. A double-tagged oligonucleotide (DTO) is employed to efficiently add two different tags to ends of DNAs to make a double-tagged DNA libraries. Also disclosed are methods to make mate pair libraries using the double-tagged oligonucleotide, and methods to make double-tagged single stranded DNA. The double-tagged DNA libraries of the invention are ready to be used on next generation sequencing machines.

主权项

1. A method of making a di-tagged single-stranded DNA from a double-stranded DNA, said method comprising the steps of:
a) providing a double-stranded double-tagged oligonucleotide (DTO) sequentially comprising a sequence tag A, a breaking site, a sequence tag B, and a single nick or gap in one of the two strands; b) ligating two ends of said double-stranded DNA to said double-tagged oligonucleotide to generate a circular DNA-DTO; c) removing the nicked/gapped strand using an exonuclease to generate a circular single-stranded DNA; d) breaking said circular single-stranded DNA at said breaking site to generate a linear di-tagged single-stranded DNA.