Molecular Cloning and Analysis of Glutathione S-Transferase Gene of Schistosoma japonicum

摘要

To construct recombinant clone of glutathlone S-transferase (GSTs) of Schistosoma japonicum (Sj) and express recombinant SjGSTs protein with his-tag that could be purified by single-step Ni2+-NTA affinity chromatography，one pair of primers was designed according to the sequence of GSTs gene. The DNA fragment of PGEX-KG GSTs gene was amplified by PCR with template PGEX-KG. Then it was cloned into vector pET28a and recombinant prokaryotic expressing vector was constructed, and corroborated through restriction enzymes map and sequencing. Identity analysis and, primary and tertiary structure of translation alignment were made using NCBI-blast and Swiss-module. Results showed that pET28a-SjGST recombinant clone was constructed successfully. The length of amplified GSTs gene was 693 bp, and has the opening reading frame (ORF) of 687 bp in length and has 99% identity with PGEX-KG-GST, Sj -GST-Mainland-26ku (SjGST-ML-26), and Sj-GST-Philippines-26ku (SjGST-PL-26). The deduced animo acid sequence consisted of 228 animo acid, PI was 6.17 and molecular weight was 26766 mw. Recombinant protein should be expressed in soluble protein form. It was transformed into E. coli BL 21 for subsequent expression of his-tagged GSTs and purification with Ni2+-NTA-sepharose affinity column.