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本研究克隆和表达了刺参原肌球蛋白(Tropomyosin,TRP)基因,进一步研究刺参再生过程中重要分子的功能。结果表明,TRP基因序列总长为1203bp,5′非翻译区为105bp,3′非翻译区为240bp,该序列包含一个855bp的开放阅读框(openreading frame,ORF),编码284个氨基酸,分子量为33.27kDa,等电点为4.56。利用Escherichiacoli对TRP进行了体外重组表达,在1mmol/LIPTG和37℃条件下诱导,能产生分子质量约为38kDa的重组蛋白,Westernblot证明重组TRP与鼠抗刺参原肌球蛋白的多克隆抗体能特异性结合。
In this study, we cloned and expressed Tropomyosin (TRP) gene to further study the function of important molecules in the process of sea cucumber regeneration. The results showed that the total length of the TRP gene was 1203 bp, the 5 ’untranslated region was 105 bp, and the 3’ untranslated region was 240 bp. The sequence contained a 855 bp open reading frame (ORF) encoding 284 amino acids with a molecular mass of 33.27 kDa, isoelectric point of 4.56. TRP was recombinantly expressed in vitro using Escherichia coli and induced at 1 mmol / L IPTG and 37 ° C to produce a recombinant protein with a molecular mass of about 38 kDa. Western blot showed that the polyclonal antibody to recombinant TRP and stubble tropomyosin Specific binding.