Construction and Identification of a cDNA Expression Library from Clonorchis sinensis Metacercaria

Objective To select stage specific-antigen gene for diagnosis and antigen gene for vaccine candidate, a cDNA expression library of Clonochis sinensis metacercaria (Csm) was constructed. Methods Total UNA from C. sinensis metacercaria was extracted and cDNA was synthesized by SMART?cDNA library protocol. Products were digested by proteinase K and Sfi I , and fractionated with CHROMA SPINT 400 column. The fractions were characterized by 1.1% agarose/EB gel electrophorese. The cDNA with size of 0.4 - 4kb were reamplified and h'gated to λ TriplEx2 vector. The Xphage packaging reaction for the ligated DNA was performed to produce an unamplified library. Then, the phage titer the percentage of the recombinants of the unamplified library were estimated. Ten phages were randomly selected and amplified using primers from vector by PCR to test the quality of obtained library. Results The tilers of unamplified and the amplified library were 7.0 × 107pfu/ml and 2.5 × 108 pfu/ml respectively. 30% of the recombinant contained an insert beyond 1kb, while the insert of 69% of the recombinants was between 500bp - 1000bp. Condution A high quality cDNA library from C. sinensis was constructed successfully.