冷盐沉淀同步分离dsRNA病毒基因组和结构多肽的方法  

A Method for Synchronously Isolating the Genome and Structure Polypeptides from dsRNA Viruses in Cold Salt Solution

作　　者：董长垣 陈晓 陈保平 严银钫 涂攀 宋麒 

出　　处：《湖北医科大学学报》2000年第4期265-267,共3页

基　　金：国家自然科学基金!资助项目 (编号 :392 70 0 31)

摘　　要：目的 :探索一条既能获得dsRNA病毒全基因组 ,又能同时获得该病毒完整结构多肽的方法。方法 :将分别感染了蓝舌病毒 (BTV)和人轮状病毒 (HRV)的培养细胞分别破碎以释出病毒后 ,结合应用差速离心和蔗糖梯度离心技术 ,获得了这两种病毒的纯化粒子 ;再用 1%的SDS裂解病毒 ,0 .2 5mol/L的KCl沉淀其结构蛋白 ,而病毒基因组悬于上清。结果 :所沉淀的蛋白用PBS(pH 7.8)适当稀释后 ,即可用作抗原 ;上清用酚∶仿抽提 1次后 ,即可用于核酸分子生物学研究。结论 :此方法为一套既能有效同步分离dsRNA病毒核酸和结构多肽等活性生物大分子 ,又能节约实验材料和时间的有效方法。In order to create the method by which the dsRNA genome and complete structure polypeptides can be isolated from the dsRNA viruses,we had the vero cell infected with the Bluetongue Virus (BTV),and the MA 104 cell infected with Human Rotavirus (HRV).The two virus infected cell strains were separately ruptured to release the virus particles from these cells;Using a difference speed centrifugation and a sucrose density gradient centrifugation,the intact viruses were obtained with ideal purity.Then,these viruses were used as the materials for synchronously isolating the structrue polypeptide and genome dsRNA with activity function.The viruses were ruptured using 1% SDS,and their structure proteins were precipitated in the presence of 0.25 mol/L cold KCl,which can be used as the antigen to study the immunology after properly diluted in PBS(pH 7.8).After the suspension was extracted one time with an equal volume of phenol/chloroform,the viral genome dsRNAs were gotten for the research of the viral molecular biology.This is a good method for synchronously isolating the genome and structure polypepetides from dsRNA viruses with biological activity.

关 键 词：蓝舌病毒 分离 提纯 轮状病毒 冷盐沉淀同步分离 

分 类 号：R373.9[医药卫生—病原生物学] R373.2[医药卫生—基础医学]