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美国白蛾几丁质脱乙酰酶1(HcCDA1)的克隆表达与酶活测定

闫晓平1, 赵丹1, 孙晓彤1, 郭巍1,2, 李少雅1, 李景2   

  1. 1. 河北农业大学生命科学学院/河北省农作物病虫害生物防治工程技术中心, 保定 071000;2. 北京农学院植物科学技术学院, 北京 102206
  • 收稿日期:2014-09-29 修回日期:1900-01-01 出版日期:2015-06-08 发布日期:2015-06-08
  • 通讯作者: 郭巍

Clone, Expression and CDA Activity of Chitin Deacetylase 1 from Hyphantria cunea (Drury)

YAN Xiaoping1, ZHAO Dan1, SUN Xiaotong1, GUO Wei1,2, LI Shaoya1, LI Jing2   

  1. 1. College of Life Sciences, Agricultural University of Hebei/Crop Diseases and Pests Biological Control Engineering Technology Center of Hebei, Baoding 071000, China;2. Plant Science and Technology College, Beijing University of Agriculture, Beijing 102206, China
  • Received:2014-09-29 Revised:1900-01-01 Online:2015-06-08 Published:2015-06-08

摘要: 利用RACE-PCR方法,扩增得到编码美国白蛾Hyphantria cunea Drury几丁质脱乙酰酶基因HcCDA1.将该基因分别在大肠杆菌BL21和毕赤酵母细胞Pichia pastoris GS115表达.经诱导表达后,Western blot分析表明HcCDA1在大肠杆菌中成功表达61.8 kD的蛋白,符合预期大小;在毕赤酵母GS115中成功表达80 kD蛋白.酶活性试验显示酵母细胞中表达的重组蛋白酶活力为1.685 U/mL.本研究利用毕赤酵母成功表达具有酶活力的重组HcCDA1蛋白,为进一步明确该蛋白的生理功能提供条件.

Abstract: HcCDA1 cDNA sequence was obtained by PCR and rapid amplification of cDNA ends (RACE). HcCDA1 was expressed in Escherichia coli BL21 and Pichia pastoris GS115, respectively. The HcCDA1 was identified to be a 61.8 kD protein in E.coli BL21 and 80 kD protein in P. pastoris GS115 by Western blot analysis. The CDA activity of recombinant HcCDA1 expressed in P. pastoris GS115 was 1.685 U/mL. The expression of recombinant HcCDA1 will facilitate further investigation of physiological functions of HcCDA1 protein.

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