Abstract: Hemocyanins are copper-containing (Cu2+) proteins in vivo of insect and play important roles not only in respiratory  but also in energy storage, antibiosis and antivirus. To investigate the structure and expression profiles of Calliptanus italicus hemocyanin subunit typeⅠ(CitHc-1), the complete cDNA (2 335 bp) of CitHc-1 was obtained by RACE method. The CitHc-1 contained an open reading frame of 2 079 bp encoding 692 amino acids. The predicted molecular weight is 79.88 kDa. Sequence alignment revealed that the CitHc-1 shared an identity of 91%~94% with hemocyanins from other Acridoidea species. The dendrograms of the arthropod hemocyanin subunit typeⅠinferred by Neighbor-joining (NJ) method showed that, Calliptanus italic hemocyanin subunit typeⅠand Locusta migratoria hemocyanin subunit typeⅠformed a subclade. In order to further study the structure and function of hemocyanin subunit typeⅠfrom Calliptanus italicus, the CitHc-1 prokaryotic expression vector was established successfully and the recombined plasmid was transfected into BL21(DE3). The result of SDS-PAGE showed that the molecular weight was consistent with the theory molecular weight (32 kDa). The results provided a theoretical basis for further analysis on the function of CitHc-1 in C. italicus.

Key words: Calliptanus italicus, hemocyanin subunitⅠ, gene clone, expression analysis, prokaryotic expression

摘要： 昆虫血蓝蛋白是在昆虫体内普遍存在的一种含Cu2+的多功能蛋白，除参与呼吸外还具有能量贮存、抗菌和抗病毒等多种生物学功能。为了研究意大利蝗（Calliptanus italicus）血蓝蛋白亚基CitHc-1的结构和表达，通过RACE技术获得了CitHc-1的全长cDNA（2 335 bp）序列，其中开放阅读框2 079 bp，编码692个氨基酸，预测蛋白分子量79.88 kDa。序列比对结果显示CitHc-1基因cDNA序列与蝗总科其他物种血蓝蛋白亚基Ⅰ的cDNA序列的相似性为91%~94%。利用MEGA的NJ法构建昆虫纲血蓝蛋白亚基Ⅰ系统发育树，结果显示意大利蝗CitHc-1与东亚飞蝗LmiHc-1血蓝蛋白遗传距离最近形成姐妹群。为了研究血蓝蛋白的结构和功能，成功构建了意大利蝗CitHc-1基因活性区域的原核表达载体。融合蛋白pEASY-E1-Hc分子量约为32 kDa，与预期值一致，为进一步分析意大利蝗C. italicus CitHc-1的功能提供了理论基础。

关键词: 意大利蝗, 血蓝蛋白亚基Ⅰ, 基因克隆, 表达分析, 原核表达