[Cloning, expression, and antibody production of mouse ISP2]

A cDNA encoding mouse implantation serine proteinase 2 ISP2 was amplified from total cDNAs of mouse uterus implantation sites on D4.5 of pregnancy by PCR, and sequenced GenBank accession No. A442918 . DNA sequencing indicated that the ISP2 cDNA had an unreported 204 bp sequence at 3' untranslated region besides the open reading frame encoding 279 amino acid residues, which was identical with literature. In order to obtain recombinant ISP2 rISP2 an expression plasmid pGEX-4T-2/ISP2 was constructed and transformed into E.coli BL21(DE3) strain. Expressed fusion protein GST-ISP2 was purified by SDS-PAGE and digested with thrombin, and the digestion mixture was subjected to SDS-PAGE again to recover rISP2. Rabbits were immunized using rISP2 as immunogen, and the polyclonal anti-ISP2 antisera were obtained. Immunohistochemical analysis using this antisera showed specific and high expression of ISP2 in mouse endometrial gland epithelium in early pregnancy.