Restriction digests of the clone give the following sizes (kb): EcoRI--8.0, 1.2; PstI--9.2; HindIII--6.3, 1.8, 1.1. The vector was designed for N-terminal tagging with GST (glutathione S transferase). The vector contains a thrombin cleavage site (LVPR/GS) immediately following the GST moiety, to allow removal of the tag following purification. The vector does not contain a stop codon. The vector was constructed by 1) amplification of the Schistosoma japonicum glutathione S-transferase gene with primers designed to flank the gene and modify the polylinker, 2) gel purification and digest of the PCR product with XhoI, 3) ligation into pREP4X cleaved with XhoI and SmaI; the SmaI site was lost during cloning, 4) Digestion with XhoI & BglII, purified GST-containing fragment was cloned into pSLF173, digested with XhoI and BglII. |
