CLONING,PROKARYOTIC EXPRESSION OF SERINE PROTEASE INHIBITOR 23 FROM AEDES AEGYPTI
10.3969/j.issn.1005-0507.2017.04.005
- VernacularTitle:埃及伊蚊丝氨酸蛋白酶抑制剂23的基因克隆表达与多克隆抗体制备
- Author:
Hui ZHAI
1
;
Zheng-Yan WANG
;
Qing-Qiao LYU
;
Jin-Zhi CHENG
;
Shi-Qi LI
;
Xi YANG
;
Zheng-Ling SHANG
;
Jia-Hong WU
Author Information
1. 贵州医科大学环境污染与疾病监控省部共建教育部重点实验室
- Keywords:
Aedes aegypti;
Serine protease inhibitors;
Cloning and expression;
Western Blot
- From:
Acta Parasitologica et Medica Entomologica Sinica
2017;24(4):230-235
- CountryChina
- Language:Chinese
-
Abstract:
Aedes aegypti serpin23 (serine protease inhibitor,23) highly specific expressed in the female mosquito salivary gland,and was shown as a major protein component of mosquito saliva with unknown physiological functions. The purpose of the present study is to clone serpin23 from Ae. aegypti Guangdong Dianbai strain and express in prokaryotic cell line for subsequent functional studies. The serpin23 coding gene was amplified with specific primers and cloned from extracted RNA of Ae. aegypti Guangdong Dianbai strains. The length of serpin23 gene was shown in length of 1197 base pairs,encoding 399 amino acids. Recombinant plasmid pET-28a ( +) -serpin23 was constructed to express serpin23 gene in E. coli BL21 (DE3). As a result,Serpin23 protein was detected in SDS-PAGE gels and purified by KCl stained technique. The Serpin23 protein expressed,about 47 kDa,was harvested in inclusions. Its isoelectric point was 6.13,containing a Serpin domain, and a functionally unknown FAINT domain. Moreover, polyclonal antibody of mouse against anti-Serpin23 protein were prepared from BALB/c mice using a standard immunizations protocol and submitted for Western Blot analysis. The antiserum form immunized BALB/c mice also recognized the expressed Serpin23 specificly. The premilinary results paved a way to explore the potential functions of serpin23 gene in salivary gland of Ae. aegypti.
