Cloning and Sequence Analysis of Myostatin(MSTN) Gene from Fujian White Rabbit
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摘要: 为掌握福建白兔负向调控骨骼肌生长发育的肌肉生长抑制素（Myostatin，MSTN）基因分子基础，应用RT-PCR和RACE技术从肌肉组织总RNA中克隆出福建白兔MSTN基因的cDNA序列并进行分析。结果表明，该基因序列全长1 551 bp（GenBank登录号：KX084386），其中5'非翻译区136 bp，3'非翻译区287 bp，1 128 bp的开放阅读框，编码375个氨基酸，MSTN蛋白在第97~768核苷酸处编码的氨基酸存在1个TGF-β前肽超家族结构域，在第841~1 125核苷酸处编码的氨基酸存在1个TGF-β家族结构域。序列分析表明：福建白兔与穴兔、人、猪、家鼠、马、牛、绵羊、鹿、狒狒、猩猩、猴的MSTN核苷酸序列同源性分别为99.9%、92.8%、94.4%、86.9%、95.2%、89.1%、91.0%、90.3%、95.9%、96.4%、95.9%。研究结果为进一步开展福建白兔MSTN基因的结构、表达以及分子育种等相关研究提供了基础数据。Abstract: In order to master the molecular basis of Myostatin(MSTN) gene that negatively regulates skeletal muscle growth and development in Fujian white rabbit, Fujian white rabbit MSTN gene cDNA was amplified and cloned from total RNA of muscle by RT-PCR and RACE(rapid amplification of cDNA ends). Sequence analysis indicated that the full cDNA (1 551 bp, Gen Bank accession number:KX084386) consisted of a 5'-terminal untranslated region (UTR) (136 bp), a 3'-terminal UTR (287 bp) and an ORF(1 128 bp). The open reading frame encoded a deduced protein with 375 amino acids residues. There were one TGF-beta propeptide superfamily domains at amino acids encoded by the 97 to 768 nucleotides and one TGF-beta family domains at amino acids encoded by the 841 to 1 125 nucleotides. Comparison of Fujian white rabbit MSTN with those of other rabbit, human, pig, mouse, horse, cattle, sheep, deer, baboons, orangutan and monkey showed that the amino acids horno1ogy were 99.9%, 92.8%, 94.4%, 86.9%, 95.2%, 89.1%, 91.0%, 90.3%, 95.9%, 96.4%, 95.9%, respectively. It provides the basic data for further research on structure function, expression regulation and molecular breeding of MSTN gene in Fujian white rabbit.
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Key words:
- Fujian white rabbit /
- MSTN /
- clone /
- sequence analysis
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图 1 产物电泳
注：A为总RNA(M：DL2000 DNA Marker，1：总RNA)；B为MSTN的同源序列(M：DL2000 DNA Marker，1:PCR产物; 2:阴性对照)；C为MSTN的3′RACE(M：DL2000 DNA Marker，1:PCR产物，2:阴性对照)；D为MSTN的5′RACE(M：DL2000 DNA Marker，1：PCR产物)；E为RACE验证(M：DL2000 DNA Marker，1：阴性对照，2：PCR产物)。
Figure 1. Products were detected by gel electrophoresis
图 2 福建白兔MSTN cDNA序列及其预测的氨基酸序列
Figure 2. Sequence of MSTN from Fujian white rabbit and its amino acid sequence
表 1 设计的引物
Table 1. Primer sequence
名称 序列(5′-3′) 长度/(mers) F1(已知序列) GTTTATGCTGATCGTGGCTG 20 R1(已知序列) CACCCACAGCGGTCTACTAC 20 F2(已知序列) CTGTGTAATGCATGCACTTG 20 F3(3′RACE) AGCAGGTCCTTGCTGTACTC 20 F4(3′RACE) CCAATTACTGCTCTGGAGAC 20 F5(3′RACE) GAGCACTCAACAGAATCACG 20 R4(5′RACE) CTTCCAAAGAGCCATCACTG 20 R6(5′RACE) CTGAACGTCGTACTGATC 18 YZF1(序列验证) CTCAAGCTGTTCATGCAT 18 YZR1(序列验证) CCAACCATTGCATGTATTC 19 
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