The Expression Of Follicle Stimulating Hormone Of Cervus Nippon

Follicle stimulating hormone (FSH) has been used widespreadly on superrovulation and artificial fertilization in propagation of cattle, sheep, pig. FSH can be obtained by several techniques, but the FSH made by the anterior pituitary from animals often had some severities matters, such as short of primary materials, lime-consuming, baborious, copy, low purity and contaminated with other hormones. Using recombinant technique, large quantity and high purity FSH can be produced and the production expenditure also can be reduced, which is important to animal husbandry.At the basis of study by Prof Guan Hongbin, this experiment focuses on the study of the Cervus nippon FSHa/βsubunit gene cloning and fusion expression.Firstly, two primers including the corresponding restriction enzyme sites of EcoR I,Xho I was designed, and FSHa/βsubunit gene segments were cloned by PCR from the original constructed vector. Then the purificated production of PCR was connected with carrier pMD18-T, and transformed into JM109. After restricted enzymes digestion by EcoR I and Xho I, the gene was inserted into the carrier pGEX-6P-2 correctly. At the end, the recombinant plasmid was built. Last, the positive clones were transformed into E.coli BL21, and then the GST-FSHα/βfusion proteins were expressed via the induction of IPTG, and detected by SDS-PAGE.The FSHa subunit was obtained by cloning, and the PCR product was about 370 bp. The sequencing result was identical with GenBank. The recombinant expression vector pGEX-6P-2-FSHa was constructed successfully, and the fusion expression protein which was induced by IPTG was analysed via SDS-PAGE. The specific protein bands appeared in relative molecular weight of 39.3 kD expectantly. The FSHβsubunit was also cloned successfully. The PCR product was about 400 bp, and the sequencing result was identical with GenBank's. And then the recombinant expression vector pGEX-6P-2-FSHβwas constructed correctly, At the end, the fusion expression protein induced by IPTG was analysed via SDS-PAGE. The specific protein bands appeared in relative molecular weight of 39.5 kD Department. Those results indicated that the FSHα/β-GST fusion protein of Cervus nippon were successfully expressed in E.coli BL21.