Sequencing library construction method

TL;DR: A sequencing library construction method disclosed by the authors in this paper comprises the following steps: step 1, further completing works of repair of a tail end of a sample DNA fragment, phosphorylation of a fragment 5' end and dA addition of fragment 3' end by an ERAT reaction system in one step; step 2, adding a connector, ligase and a ligase buffer solution into the ERAT repair sample obtained in the step 1 and retaining a small fragment DNA in the sample according to the needs of a library; step 3, purifying and recovering a connection

Abstract: A sequencing library construction method disclosed by the invention comprises the following steps: step 1, further completing works of repair of a tail end of a sample DNA fragment, phosphorylation of a fragment 5' end and dA addition of a fragment 3' end by an ERAT reaction system in one step; step 2, adding a connector, ligase and a ligase buffer solution into the ERAT repair sample obtained in the step 1, and retaining a small fragment DNA in the sample according to the needs of a library; step 3, purifying and recovering a connection product by 1x-2x magnetic beads with the registered trademark of AMPure XP, wherein the fragment is screened through adjusting the ratio of magnetic beads and other aspects according to the needs; step 4, carrying out PCR reaction for 7-11 cycles by using high-fidelity enzymes, and amplifying the connection product; and step 5, purifying and recovering the PCR product with the magnetic beads with the registered trademark of AMPure XP, and carrying out quality control detection to complete library construction. The suitable reaction system and fewer library construction steps are used, and the cost of library construction can be effectively reduced.