中国三黄鸡可溶性B淋巴细胞刺激因子基因的克隆、表达和活性研究  

Cloning, expression and characterization of soluble BAFF in Chinese 3-yellow chicken

作　　者：陈舒劼[1] 刁振宇[1] 张超[1] 张双全[1] 

机构地区：[1]南京师范大学生命科学学院,江苏省分子医学生物技术重点实验室,南京210097

出　　处：《动物学报》2007年第3期531-536,共6页ACTA ZOOLOGICA SINICA

基　　金：江苏省科技厅自然科学基金(批准号BK2005140);南京师范大学科技创新基金(批准号184070H809)联合资助~~

摘　　要：通过逆转录-聚合酶链式反应,从中国三黄鸡脾组织细胞中扩增得到459bp的鸡可溶性B淋巴细胞刺激因子(CycsBAFF)基因片段,将其克隆入原核表达载体pET-28a,转化大肠杆菌BL21(DE3)后高效表达了带有His6标签的包涵体形式重组蛋白,通过聚丙烯酰胺凝胶电泳及免疫转印检测鉴定。表达产物经Ni2+-NTA纯化后,经透析复性得到活性目的蛋白,通过单独刺激以及与PMA共刺激体外培养的鸡法氏囊B细胞,均有明显的促法氏囊B细胞存活的效应。459 bp Chinese 3-Yellow chicken soluble BAFF （CycsBAFF） cDNA was amphfied by reverse transcription polymerase chain reaction （RT-PCR） method using total RNA isolated from chicken spleen as template, and then inserted into pET-28a vector. After the recombinant vector was transformed into E.coli BI21 （DE3） which was then induced with Isopropyl-β-Dthiogalactopyanoside （IPTG）, the recombinant protein with polyhistidine tag （Hiss-Tag） was expressed with high efficiency as inclusion bodies and identified by SodiumDedecylSulfate-Polyacrylamide Gel Electrophoresis （SDS-PAGE） and Western blot. After the target protein was purified by Ni2. -nitrilotriacetic acid （Ni^2＋ -NTA） and refolded by dialysis, the bioactivity of recombinant CycsBAFF was analyzed by stimulating and co-stimulating chicken bursal B cells with Phorbol 12-Myristate 13-Acetate （PMA）, and both suggest obvious effects of promoting chicken bursal B cell survival [Acta Zoologica Sinica 53 （3） ： 531 - 536, 2007]

关 键 词：鸡可溶性B淋巴细胞刺激因子 基因 克隆 表达 法氏囊B细胞 

分 类 号：S852.4[农业科学—基础兽医学]