关键词: 和田羊, 卡拉库尔羊, 多浪羊, 肌生成抑制素, CHO细胞, 绿色荧光蛋白

Abstract: This work aims to construct the fusion expression plasmids of myostatin gene（MSTN）from 4 local varieties（lines）sheep，namely plain Hetian sheep（HP），mountain Hetian sheep（HS），Carla Kul sheep（KL），and Duolang sheep（DL）in the southern Xinjiang with enhanced green fluorescent protein（EGFP），and the expression of MSTN in eukaryotic cells. Total RNA was extracted from the skeletal muscle in 4 varieties of sheep，cDNA was acquired by reverse transcription，and MSTN amplified by PCR was ligated with pMD18-T vector to construct recombinant plasmid and sequenced. Then MSTN（HP，HS，DL，and KL）gene was inserted into the fusion expression vector pEGFP-N1，the recombinant plasmid of pEGFP-N1-MSTN in a eukaryotic fusion expression was constructed. The plasmid with over 80% cells covered was transfected into CHO by liposome，the green fluorescent of fusion protein of EGFP and MSTN in CHO cells was observed. The results showed that the ORF sequence of MSTN gene in any of HP，HS，KL，and DL was 1 125 bp（excluding stop codon），encoding 375 amino acids. Comparing MSTN sequences in GenBank，there was differences in HS and HP；and 2 nucleotide variation in KL caused 2 amino acid mutations；and 6 nucleotide differences in DL resulted in 4 amino acid variations. In conclusion，the recombinant plasmid for EFGP and MSTN co-expression was constructed and expressed in eukaryotic CHO cells.

Key words: hetian sheep, carla kul sheep, duolang sheep, myostatin, CHO cell, green fluorescent protein