| The cDNA sequence encoding Cyprinus carpio insulin-like growth factor-â… (IGF-â… ) was cloned from the fresh Cyprinus carpio liver by means of using thespecific primers based on the reported sequence in GenBank (D83272). The cDNAwas inserted into to the pMD18-T plasmid, then identified to be IGF-â… with enzymedigestion and sequence identification. The full length of the cloned cDNA, was 538bp with an ORF composed of 486 nucleosides, which encodes 162 amino acids. Italso showed there was high homology between the Cyprinus carpio IGF-â… and otherIGF-â… sequence published from the GenBank.The nucleosides was was an identity of94.4%-99.0%, and the identity of deduced amino acids was 94.0-97.3%. Meanwhile,the Hydrophobicity and surface probability analysis of IGF-â… protein were the sameas the feature of IGF-â… protein.By the technology of DNA recombination, the IGF-â… cDNA was cloned intoProkaryotic Expression plasmids pBV220 vector, and was transformed into E.coli.The recombinant plasmid was induced to expression by different temperature. Theresults of SDS-PAGE showed that the expression products were about 7KD andreached their expression peaks about 6 hours after inducing.The result also showedthat the expression quantity of fusion protein had 15.0% ratio of total thalline protein,while the E.coli without inducing by temperature didn't express. The expressednon-inclusion body IGF-â… were washed,then the biological activity were detected byMTT. The results indicated that the recombinant IGF-â… proteins have the high abilityto supply the proliferation of CIK cells by t test, and have higher biological activity.The IGF-â… was cloned firstly in China and the IGF-â… protein were prokaryoticexpressed in this experiment. The protein could be used for weight promoting in vivoor in vitro; meanwhile, the further research of the IGF-â… is basic for the mechanismthat IGF-â… acted in molecule controlling, signal conduction, embryonic development,reproduction and immunization.
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