火菇素cDNA的初步免疫学筛选  

Purification of flammulin and preliminary immunoscreening of it's cDNA

作　　者：周凯松[1] 薛久刚[1] 黄芳[1] 张晗星[1] 张长铠[1] 

机构地区：[1]山东大学微生物技术国家重点实验室,山东济南250100

出　　处：《山东大学学报（理学版）》2004年第2期120-124,共5页Journal of Shandong University(Natural Science)

基　　金：山东省自然科学基金重点资助项目 (NOZ2 0 0 2D0 1) ;国家自然科学基金资助项目 (NO3 0 3 70 0 14 )

摘　　要：火菇素是一种具有抗癌活性的简单蛋白 ,从金针菇中分离纯化火菇素的方法如下 :金针菇子实体经乙酸液浸提、硫酸铵和乙醇分级沉淀后 ,通过DEAE SephadexA 5 0除去色素和部分杂蛋白 ,最后经CM SephadexC 5 0离子交换柱层析分离 ,得到了电泳均一的纯化蛋白 .以火菇素为抗原制备的抗血清为探针对构建的金针菇cDNA表达文库进行免疫印记筛选 ,对获得的阳性克隆进行PCR、酶切以及在大肠杆菌中诱导表达等初步鉴定 ,为进一步确证、克隆火菇素基因打下了基础 .Flammulin,an anti-tumor protein,was purified from the aqueous extract of basidiomes of Flammulina Velutipes . The purification process included ammonium sulfate and ethanol fraction,DEAE-Sephadex A-50 and CM-Sephadex C-50 ion-exchange chromatography. Flammulin was purified to electrophoretic homogeneity. the cDNA library constructed from Flammulina velutipes was immunoscreened using antibodies against purified flammulin,a candidate cDNA related with flammulin gene was primarily determinated by PCR,restriction mappling analysis and SDS-PAGE analysis. This will serve as a basis for further screening and cloning flammulin gene.

关 键 词：火菇素 金针菇 CDNA 表达文库 免疫筛选 

分 类 号：Q75[生物学—分子生物学]