Construction of directional cDNA library from mixed human 1Hepatocellular Carcinoma tissue

[Objective] To quickly construct a directional cDNA library from mixed human hepatocellular carcinoma tissues. [Methods] The total RNA was separated from mixed human HCC (hepatocellular carcinoma) tissues, then the first strand cDNA was synthesized with oligo (dT) primer containing Sfi I-digested sites before the double-strand cDNA was amplified through LD-PCR (Long-distance PCR) by SMART technology. The double-strand cDNA was digested by Sfi I ( IA IB) restriction enzyme before cDNA size fractionation, the double-strand cDNA fractionated was ligated into the λ Trip Iex-vector and packaged in vitro. [Results] The unamplified mixed human hepatocellular carcinoma tissues cDNA library consisted of 3.96 ×106 independent clones with recombinant clones more than 93%. The titer of the amplified cDNA library was 3.92×109 pfu/ ml and the average size of the recombinants insert 1. 3 kb. [Conclusion] The quality of the constructed mixed human HCC tissues cDNA library is excellent and helpful to screen HCC specific antigen.