Construction Of Recombinant Adenovirus Of Mink Growth Hormone Gene

As everyone knows that the mink is a kind of fur animal which has high economy value.By the research, the size and the quality of the skin are not only about with the different species,the nourishment and feeding level,but also closely related with the physiology condition of the mink.The mink growth hormone(mGH) is a kind of protein hormone which excreted from the brain adenohypophysis cells;it has the extensive physiology function.If the growth hormone was injected to the animals' bodies,it could obviously increase their growing speed,and increase bodies' length,raising the furrieries' quality.But the natural growth hormone yield is very tiny.It's hard to wide-ranging content need,nevertheless.The development of genetic engineering technique and virus vector carry resolves this problem effectively.In this paper,using adenovirus vector to produce genetically engineered growth hormone MinkFirstly,as the results on animal experiment of mink with the growth hormone gene eukaryotic expression vector,we transient transfected the mink growth hormone gene eukaryotic expression vector pCMTmGH in BHK cells,and get the culture fluid by SDS-PAGE detection after 48 hours,the result shows that vicinity of 22 KD has a purpose strap.It preliminary shows that the vector can be expressed in eukaryotic cells.Provide the basis for useing the double-promoter to construct the adenovirus shuttle plasmid about mink growth hormone gene.Secondly,according to the requirement,we add HindⅢand KpnⅠrestriction site at the end of specific PCR primers of mink growth hormone gene,and then linked the gene to the pEasy-T vector,constructed pmGH cloning vector.Sequenced results show that sequence has been successfully extended to Mink growth hormone gene and the two restriction sites had added to the two ends of mink growth hormone gene.We cut down the dual-promoter of eukaryotic expression vector of pCMTmGH and link with the pΔEMTCD large fragment to constructed pΔECMTCD,and then use HindⅢand KpnⅠcut off the pGHcDNA of pΔECMTCD.Even use HindⅢand KpnⅠrestriction sites to link the mink growth hormone gene with the large fragment of pΔECMTCD.PΔECMTmGH is constructed successfully by sequenced results.The last,Used the human adenovirus vector,we co-transfected the shuttle plasmid pΔECMTmGH which contained the mink growth hormone gene and the packaged vector pBHGE3 which contained the DNA fragment of adenovirus genome in 293FT cells,let them homologous recombination in 293FT cells.By the identification of electron microscopy and PCR,we have constructed the recombinant adenovirus of mink growth hormone gene under the control of CMV and MT promoter successfully.