Expression Of The Porcine Growth Hormone In Eukaryotic Cell And Microencapsulated

The growth hormone is a peptide hormone which is secreted and synthesized by acidophilic cell of anterior pituitary, it can regulate growth and development. GH has widly effect of accommodation for the growth of rhagiocrine cell and substance metabolism, it can promote growth for bone,cartilage, including other tissue, otherwise, it may stimulate protein and collagenic synthsis. its single chain is constructed by 191 amino acids, its gene length is 2231bp and the molecular mass is 2.2Kda. In practice, if supply exogenous pGH it can protein synthesis and fat moblizition. So growth velocity, forage reward and lean meat rate are elevated significantly, it posses visibly economic returns and social effects.It is reported in empirical study that the exogenous pGH can promote growth, but there are a lot of restriction, so practical application don't evolve. The key restrict factor is the route of administration, it is also a investigative focus. Now, main methods of dosage are injection with many times and the skill of slow-release in vivo embedding, but, frequently injection and operative procedure are not only trouble but also causing stress reaction for pig. The optimal way is disposably administration, in order to secete pGH lastly, stably and long. The empiriment though the technology genetic recombination. The cell line which can express stably is screened, in order to make pGH survive and secrete activitivly, so the ccell is microencapsulated.The RNA is extracted from pituitary gland, then cDNA is obtained by reverse transcription and a fragment of gene that is 678bp is obtained through analysis. The eukaryotic expression vector(pcDNA3.1-pGH) which is transfected in Vero cell is constructed. Exploitating the characteristic of Neo gene, the lowest lethal concentration is definited and the cell which can stably express pGH is got to screen with G418. The pGH gene is detected to present by the way of RT-PCR, according to this, it is proved that transfection and screening succeed. SDS-PAGE indicated that there was a conspicuous strap in 22Kda conformiting typical pGH sample. The expression is up to 12ng?106?12h in Vero cell with ELISA kit of porcine growth hormone.The microencapsulation cell can validly overcome these difficulty, it offers a new way to make use of manufacture. As a valid tool for cellular transplant, the preparating process is simple, the volume of dead space is small and the provision of nutrition is profit, so, the microencapsulation cell can react fast for the accommodative factor, for example, the glucose in host.The microcapsule is thin but transplanting cell can not reveal because its mechanical strength is good. It difficultly cause host to evoke immune reaction owning to atoxic materials, small immunogenicity and fine biocompatibility. So microencapsulation have action of immunoprotection and immunoisolate.All the merits of microcapsule are used, the cell in exponential phase of growth is microencapsulated. When the cell density is adjusted for 3×106?mL, the mixture of cells and Alginic Acid was manufactured with correlated equipment into colloidal particle, the membrane of microcapsule was polylysine.The microcapsule was in suspension in Alginic Acid,after the acidification of Citric Acid , the microcapsule changes into APA collocystis. The cytoactive was detected with MTT in the collocystis, we discovers the cytoactive to descend, and the microencapulation achieve exponential phase of growth, otherwise, up to57d, the cell growth mildly. The results of glucose and lactic acid display the consumption of glucose is stably up to 57d, and the creation of lactic acid is corresponding to the consumption of glucose. The clear supernatant liquid is detected, the result show that pGH can express lastly and stably.The result showed that, with gene recombination,we can get mammalian cell line, which persists in secreting pGH stably, and microencapsulation cells grow and secrete pGH actively.Our experiment complete basic section of transplantation for microcapsule in pigs,and offer a convenient and efficient way for the application of pGH to cultivation.