|本期目录/Table of Contents|

[1]张 婵,朱明星,杨风琴,等.结核分枝杆菌宁夏分离株pstS1基因在毕赤酵母中的表达[J].宁夏医科大学学报,2013,(08):860-863.
 ZHANG Chan,ZHU Mingxing,YANG Fengqin,et al.Expression of Mycobacterium Tuberculosis pstS1Gene Isolated from Ningxia in Pichia Pastoris[J].Ningxia Medical University,2013,(08):860-863.
点击复制

结核分枝杆菌宁夏分离株pstS1基因在毕赤酵母中的表达(PDF)
分享到:

《宁夏医科大学学报》[ISSN:1005-8486/CN:64-1029/R]

卷:
期数:
2013年08期
页码:
860-863
栏目:
论著
出版日期:
2013-08-20

文章信息/Info

Title:
Expression of Mycobacterium Tuberculosis pstS1Gene Isolated from Ningxia in Pichia Pastoris
作者:
张 婵1 朱明星2 杨风琴3 王秀青3
1.宁夏人民医院神经内科,银川 750021; 2.宁夏医科大学实验动物中心,银川 750004; 3.宁夏医科大学检验学院,银川 750004
Author(s):
ZHANG Chan1 ZHU Mingxing2 YANG Fengqin3 WANG Xiuqing3
1.Department of Neurology,Ningxia People's Hospital, Yinchuan 750021; 2.Experimental Animal Centre, Ningxia Medical University,Yinchuan 750004; 3.School of Clinical Examination, Ningxia Medical University,Yinchuan 750004
关键词:
结核分枝杆菌 pstS1基因 毕赤酵母 表达
Keywords:
mycobacterium tuberculosis gene of pstS1 pichia pastois expression
分类号:
Q343.1+2
DOI:
-
文献标志码:
A
摘要:
目的 亚克隆结核分枝杆菌宁夏分离株pstS1基因,构建pPICZα-A-S1表达载体,并在毕赤酵母表达系统中分泌表达,为pstS1基因所表达的蛋白功能的研究奠定基础。方法 PCR方法亚克隆结核分枝杆菌宁夏分离株pstS1基因片段,构建毕赤酵母表达载体pPICZα-A-S1,重组质粒线性化之后电转化到在毕赤酵母表达菌株SMD1168感受态细胞中,挑选阳性克隆菌株并用甲醇进行诱导表达,表达产物通过SDS-PAGE进行鉴定。结果 构建了结核分枝杆菌pPICZα-A-pstS1重组质粒,以甲醇进行诱导,经过SDS-PAGE电泳进行分析,结果显示pstS1基因表达出了大约为38kD的蛋白,经过Western blot分析,分泌的蛋白与结核分支杆菌38KD抗血清能够特异性的识别。结论 结核分枝杆菌pstS1基因可以在毕赤酵母中进行很好的分泌表达,为以后pstS1基因的表达产物在疫苗研制、诊断血清研究等方面的应用奠定了基础。
Abstract:
Objective To sub-clone the gene of pstS1 which was the isolated Mycobacterium tuberculosis strains in Ningxia, to construct pPICZα-A-S1 expression vector and express in Pichia pastoris expression system and to lay a foundation for the study of the protein's function which was expressed by pstS1. Methods The gene of Ningxia Mycobacterium tuberculosis isolated strains pstS1 was sub-cloned with PCR and Pichia expression vector pPICZα-A-S1 was constructed.The recombinant plasmid after linearization was transformed into competent cells of the Pichia pastoris expression strain SMD1168. The positive cloning strain was selected and induced with methanol. The expressed product was identified by SDS-PAGE. Results Mycobacterium tuberculos pPICZα-A-pstS1 recombinant plasmid was constructed and induced with methanol. After analysis by SDS-PAGE electrophoresis, the results showed the protein expressed by pstS1 was approximate 38KD.Western blot analysis proved the specific binding capacity of secreted proteins to Mycobacterium tuberculosis 38KD serum. Conclusion Mycobacterium tuberculosis pstS1 gene can be well secreted and expressed in Pichia pastoris. It lay a foundation for the future application of pstS1 gene product in vaccine development, diagnostic serum studies and other aspects.

参考文献/References:

[1] 杨桂茂,朱明星,张爱君,等.结核分枝杆菌宁夏分离株pstS1基因的扩增及生物信息学分析[J].宁夏医科大学学报,2010,9(32):960-962.
[2] 萨姆布鲁克J,弗里奇EF,曼尼阿蒂斯.分子克隆实验指南[M].2版//金冬燕,黎孟枫,译.北京:科学出版社,1987.
[3] 王秀青,张素芳,苏春霞,等.杂合抗菌肽CecA_Mag 的人工合成及其在Pichia pastoris 中的分泌表达[J].微生物学报,2004,47(1):75-78.
[4] 剧 海,粱东春,郭刚,等.用于PCR实验的毕赤酵母基因组DNA制备方法的比较[J].天津医药,2003,31(5):270-272.
[5] Lyashchenko K, Colangeli R, Houde M, et al. Heterogeneous antibody responses in tuberculosis [J]. Infect Immun, 1998, 66(8): 3936 -3940.
[6] Weldingh K, Rosenkrands I, Okkels LM, et al. Assessing the Serodiagnostic Potential of 35 Mycobacterium tuberculosis Proteins and Identification of Four Novel Serological Antigens [J].Clin Microbiol, 2005,43(1): 565-572.

相似文献/References:

[1]石华,戈朝晖,贾伟,等.结核分枝杆菌分泌蛋白MPB64表达纯化[J].宁夏医科大学学报,2011,(08):704.
 SHI Hua,GE Zhao-hui,JIA Wei,et al.[J].Ningxia Medical University,2011,(08):704.
[2]刘 阳,曹秀琴,杨志伟.嗜肺军团菌flaA部分基因的克隆及其原核表达[J].宁夏医科大学学报,2011,(03):206.
 LIU Yang,CAO Xiu-qin,YANG Zhi-wei.Expression and Cloning of Partial flaA Gene of Legionella Pneumophila in Prokaryotic Cell[J].Ningxia Medical University,2011,(08):206.
[3]马江涛,詹军,刘在新,等.杆状病毒表达系统的发展[J].宁夏医科大学学报,2009,(02):263.
[4]杨桂茂,朱明星,张爱君,等.结核分枝杆菌宁夏分离株pst S1基因的扩增及生物信息学分析[J].宁夏医科大学学报,2010,(09):960.
 YANG Gui-mao,ZHU Ming-xing,ZHANG Ai-jun,et al.Amplification and Bioinformatic Analysis of the pst S1 Gene from Ningxia Isolated Mycobacterium Tuberculosis[J].Ningxia Medical University,2010,(08):960.
[5]丁淑琴,王 洁,张 焱,等.结核分枝杆菌phoS2基因的克隆及序列分析[J].宁夏医科大学学报,2010,(06):669.
 DING Shu-qin,WANG Jie,ZHANG Yan,et al.Cloning and Sequence Analysis of the PhoS2 Gene from Mycobacterium Tuberculosis [J].Ningxia Medical University,2010,(08):669.
[6]朱佳佳,刘宏鹏,张爱君,等.结核分枝杆菌ESAT-6诊断抗原基因的克隆及同源性分析[J].宁夏医科大学学报,2010,(03):339.
 ZHU Jia-jia,LIU Hong-peng,ZHANG Ai-jun,et al.Cloning and Homologous Analysis of the ESAT-6 Gene from Mycobacterium Tuberculosis [J].Ningxia Medical University,2010,(08):339.
[7]刘胜泉,曹秀琴,杨志伟.嗜肺军团菌pip基因原核质粒的构建及其表达[J].宁夏医科大学学报,2012,(01):12.
 LIU Sheng-quan,CAO Xiu-qin,YANG Zhi-wei.Construction and Expression of Pip Gene of Legionella Pneumophila in Prokaryotic Cell[J].Ningxia Medical University,2012,(08):12.
[8]朱明星,杨桂茂,孙惠敏,等.结核分枝杆菌宁夏分离株MPT64基因的生物信息学分析[J].宁夏医科大学学报,2013,(07):778.
 ZHU Ming-xing,YANG Gui-mao,SUN Hui-min,et al.The Bioinformatic Analysis of the MPT64 Gene of Mycobacterium Tuberculosis Isolated in Ningxia[J].Ningxia Medical University,2013,(08):778.
[9]王 博,丁 晓,颜 贝,等.结核分枝杆菌Ag85B、TB10.4及Ag85B-TB10.4融合基因 的克隆表达及生物信息学分析[J].宁夏医科大学学报,2013,(07):738.
 WANG Bo,DING Xiao,YAN Bei,et al.Cloning, Expression and Bioinformatics of the Ag85B,TB10.4 and Ag85B-TB10.4 Fusion Gene in Mycobacterium Tuberculosis[J].Ningxia Medical University,2013,(08):738.
[10]张瑞芹,汤建中,贾 伟,等.结核分枝杆菌优势蛋白PPE37重组载体的构建及原核表达[J].宁夏医科大学学报,2013,(03):240.
 ZHANG Rui-qin,TANG Jian-zhong,JIA Wei,et al.The Construction and Prokaryotic Expression of Mycobacterium Tuberculosis Dominant Protein PPE37 Recombinant Vector[J].Ningxia Medical University,2013,(08):240.

备注/Memo

备注/Memo:
收稿日期:2013-03-19 基金项目:2008年宁夏高等学校科学研究项目; 宁夏医科大学特殊人才启动项目(XT200907) 作者简介:张婵(1983-),女,护师。E-mail:zhangchan83@yhaoo.com.cn 通信作者:王秀青(1979-),女,博士,副教授。 E-mail:xiuqingwang1979@yahoo.com.cn
更新日期/Last Update: 2013-08-20