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具生物学活性的重组鸡Leptin的制备
  • 摘要

    将鸡Leptin成熟肽的cDNA片段插入表达载体pRSET A的Nhe Ⅰ和Hind Ⅲ两位点之间,构建重组表达质粒pLep-SCAU2并转化大肠杆菌(Escherichia coli)菌株BL21(DE3),将重组菌在含氨苄青霉素100μg/mL的LB培养基中培养至OD600大于0.6之后,经IPTG 0.05mol/L诱导表达出分子量约为17.5 kD的含6×His标记的重组鸡Leptin,诱导4 h后表达量达最高,占总菌体蛋白的25%左右,且以包涵体形式存在.该重组鸡Leptin经50%Ni-NTA树脂纯化和透析复性折叠后分离出可溶性部分.对35日龄的矮脚黄鸡腹腔注射该可溶性重组鸡Leptin(2 mg/kg体重);注射后60 min内的采食量与对照组差异不显著(P>0.05),但在注射后80~120min都显著低于对照组(P<0.05).Leptin处理公鸡的采食量在注射2 h后逐渐上升并在5.5 h时与对照组公鸡相同.但Leptin处理母鸡的采食量持续受到抑制,在注射后5.5 h时仍然显著低于对照组母鸡(P<0.01).表明所表达的重组鸡Leptin能显著抑制鸡的采食量(母鸡比公鸡效果更为明显),证明所制备的重组鸡Leptin融合蛋白具有生物学活性.

  • 作者

    刘颖  刘志  施振旦  李孝伟  钟子穗  LIU Ying  LIU Zhi  SHI Zhen-dan  LI Xiao-wei  ZHONG Zi-sui 

  • 作者单位

    华南农业大学动物科学学院,广州,510642

  • 刊期

    2006年5期 ISTIC PKU

  • 关键词

    鸡Leptin融合蛋白  表达和纯化  复性  生物活性 

参考文献
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