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

Abstract

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.