A hybridization-based approach for quantitative and low-bias single-stranded DNA ligation

Anal Biochem. 2013 Apr 15;435(2):181-6. doi: 10.1016/j.ab.2013.01.008. Epub 2013 Jan 31.


Single-stranded DNA (ssDNA) ligation is a crucial step in many biochemical assays. Efficient ways of carrying out this reaction, however, are lacking. We show here that existing ssDNA ligation methods suffer from slow kinetics, poor yield, and severe nucleotide preference. To resolve these issues, we introduce a hybridization-based strategy that provides efficient and low-bias ligation of ssDNA. Our method uses a hairpin DNA to hybridize to any incoming acceptor ssDNA with low bias, with ligation of these strands mediated by T4 DNA ligase. This technique potentially can be applied in protocols that require ligation of ssDNA, including ligation-mediated polymerase chain reaction (LMPCR) and complementary DNA (cDNA) library construction.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • DNA Ligases / metabolism
  • DNA, Single-Stranded / analysis*
  • Gene Library
  • Inverted Repeat Sequences
  • Kinetics
  • Nucleic Acid Hybridization*
  • Polymerase Chain Reaction


  • DNA, Single-Stranded
  • DNA Ligases