Gene Cloning, Expression And Activities Analysis Of Follicle Stimulating Hormone Of Cervus Nippon

Follicle stimulating hormone (FSH) had been used widespreadly on superrovulation and artificial fertilization in propagation of cattle,sheep.FSH can be obtained by several techniques.One of them was by purifying urine from menopausal women and extracted FSH from the anterior pituitary of pig,cattle.sheep or FSH was made by using recombinant technique.But the FSH made by urine or the anterior pituitary often had some severities matters^uch as short of primary materials,time-consuming,baborious,copy,low purity and contaminated with other hormones.Those problems can be solved by using recombinant technique.The rFSH which was made by using recombinant technique,which it had some superiorities,for example providing enough primary materials,simplified generative technics, decreased generative cost,not contaminated with other hormones and differences among batches were minimized,no antibody reacrion.The gene cloning,expression and bioactivities of FSH of Cervus nippon was made firstly by RT-PCR,SDS- PAGE,Western blotjladioimmunoassy et al techniques.The results were showed as follows:1. The nucleotide sequence which encoding Cervus nippon FSH-a subunit was composed of 363 nucleotides.lt was composed of 120 amino acids.lt was directly submitted to GenBank and accepted the number as AY066018. The nucleotide sequence of Cervus nippon FSH-a subunit was compared with other animals.lt had the highest homology with goat,which reached 99%.There were only 3 nucleotides difference. It had lowest homology in nucleotide sequence with guinea pig,Callithrix jacchus, which is 83%-84%. There are 57*59nucleotides difference. It had the highest homology of amino acid sequence of Cervus nippon with goat, which reached 99%. There was only 1 amino acid difference between Cervus nippon and goat It had higher homology among bovine, sheep and buffalo, which reached 92%-93%.There were 3-4 amino acids difference among them. It was found that the nucleotide sequence of the a-subunit in these animals were highly conservative.2. The nucleotide sequence which encoding Cervus nippon FSH-b subunit was composed of 390 nucleotides. It was composed of 129 amino acids.It was submitted to GenBank and accepted as AY 156688. The nucleotide sequence of Cervus nippon FSH-b subunit was compared withotheranimals.lt had the highest homology among Cervus nippon, sheep, goat and bovine in nucleotide sequence, which reached 92%-93%. There were 24-30 nucleotides difference among them. It had lower homology in nucleotide sequence among Rattus norvegtcusjatfhodopus sungorus,which it was only 81%-82%. There were 71-74 nucleotides difference among them.Its amino acid sequence was compared with some other animals.The result showed that it had the highest homology with sheep,goat,bovine,which reached 92%-93%.There were 9-10 amino acids difference among them. It had lowerhomology in amino acid with Coturnix japamca,Nippoma nippon, its homology was 60%-65%.There were 45-52 amino acids difference among them.3. The recombinant FSH stimulated the growth of Kunming mouse's follicle.It stimulated the secondary follicle to mature follicle.The recombinant FSH stimulated the growth of Kunming mouse's uterus and ovary.The product of recombinant FSH of Cervus nippon raised the content of E2 in mouse's serum, but it had different effect on different body.Doctor: Guan Hongbin Major Basic Veterinary Science Supervisor: Professor Li Qingzhang...