Recombinant Construction Of Grass Carp Growth Hormone Gene And Analysis Of The Effect Of Transducing Fertilized Eggs Of Trout Trout

In the early stage of this study,the backcross generation from Ctenopharyngodon idella???×Squaliobarbus Curriculus???F₁ were obtained by hybridization and morphological analysis was carried out.The results showed the growth performance of backcrosses were significantly higher than that of the parents.And the head shape and eye color of backcrosses were similar with F₀ and F₁,respectively.Notably,the number of scales below lateral line was significantly different from both parents.According to the heterosis index?iH?,80.00%of qualitative traits had a higher performance than that of both parents.We had reason to believe that the growth character difference of the backcrosses is closely related to the change of genetic material.Therefore,we designed this transgenic experiment.By restructuring and molecular cloning technology in the transgenic experiment,the coding sequence of GH gene of Crass carp and the control of carp?-actin gene promoter was cloned under the pTgf2-EFla-eGFP which has been enzyme digest.The recombinant was then injected into the fertilized eggs of barbel chub?Squaliobarbus curriculus?by the method of micro-injection.It obtained mass of all-fish transgenic barbel chubs.This experiment provided research materials and ideas for fast growth rate and disease-resisted breeding for barbell chub.In order to amplify the open reading frame?ORF?of Grass carp GH gene and the coding sequence of Carp?-actin gene,a pair of Primers were Designed according to the sequence of Grass carp GH gene expression vector provided by Researcher Hu Wei.The cloned sequence was connected with pTgf2-EFla-eGFP cloning Vector and transformed into Escherichia coli DH5a.Then extract the plasmid from the bacteria and treated with XhoI and BgI?.It verified the correctness of the recombinant,then an all-fish GH gene recombinant?Transposon-Tgf2+GH gene of Crass carp+Common Carp beta-actin Promoter?was constructed,named pCAgcGH-Tgf2-L-R.The recombinant gene was extensively prepared.The recombinant and the auxiliary plasmid?pCS2-CMV-gfTP?were diluted and then mixed with 1:1.The mixture was transferred into fertilized zygotes of barbell chub by micro-injection.A group of all-fish transgenic barbell chubs were obtained.According to the sequence of Grass carp GH gene provided by academician Zhuzuoyan,a pair of Primers were Designed.Polymerase chain reaction?PCR?technology were carried out to verified transgenic barbel chubs.It turned out that there is a bright band that is consistent with positive?plasmids?,and in the comparision with the growth rate of the common grass carp and barbel chub,the growth rate of transgenic barbell chub was nearly 4 times than the normal barbell chub,but less than the grass carp.The results showed that all-fish GH gene had integrated into the genome of barbel chub,and it affected the growth rate of transgenic barbell chub to a certain extent.