斑馬魚肌肉倍增基因重組蛋白質之表現條件探討

The Expression of Recombinante Zebrafish Myostatin Protein

10.29770/JTCMT.200903.0002

李秋月(Chiou-Yueh Lee)

斑馬魚 ； 肌肉倍增基因 ； 重組蛋白質之表現 ； Zebrafish ； Myostatin ； recombination protein

台北海洋技術學院學報

2卷1期（2009 / 03 / 01）

11 - 21

繁體中文

本實驗利用RT-PCR的方法合成肌肉倍增基因，並確認為目標基因後，將其構築到pET-20(+)的表現載體上，並利用不同條件探討MSTN基因最佳表現條件。經實驗結果取得一段541bp182個胺基酸之MSTNII重組蛋白的選殖株。在不同IPTG濃度下0.1~1.0mM及不同誘導時間2~28小時作用時，重組蛋白最佳表現條件為以IPTG0.4mM誘導8小時後有最佳表現。經大量表現發現MSTN重組蛋白主要在菌體的沈澱物中，經鎳管柱分離後可得到純化的MSTN重組蛋白質。

In this study, we designed specific primers, and amplified the MSTNIT partial fragment by RT-PCR. The amplified DNA fragment was inserted into pET-20b(+) vector. The obtained MSTNII expression plasmid was transformed into E. co/i to express recombinant MSTNII protein with 182 amino acid residues. The optimal expression condition was 0.4 mM TPTG and 8 hr induced. The IPTG-induced insoluble recombinant protein was dissolved in a denatured solution and purified by a nickel-nitrilotriacetic acid (Ni-NTA) agarose column affinity chromatography via 6-histidines on its C-terminal.

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