大麦黄花叶病毒(BaYMV)的提纯

 本文提出了一个改进了的大麦黄花叶病毒提纯方法。病大麦叶片在高浓度(0.5M)磷酸钾缓冲液pH7.0(内含0.1% ME,0.01M EDTA)中匀浆,经1/4体积四氯化碳澄清后,病汁液用6% PEG,3% NaCl和1% TritonX-100混合物沉淀,高浓度(0.5M)同种缓冲液(含0.5M尿素,0.1% ME和0.01M EDTA)悬浮,接着进行20%蔗糖垫(含0.3% Triton X-100)超迷离心去除寄主细胞成分,10-40%蔗糖密度梯度离心进一步纯化。所获得的BaYMv提纯制剂A260/A280,A260/A240比值分别为1.20和1.04,纯病毒产量约为5.5-8.0mg/kg病叶。提纯病毒制剂在电镜下看不到有寄主杂质存在。

PURIFICATION OF BARLEY YELLOW MOSAIC VIRUS (BaYMV)

A purification procedure of barley yellow mosaic virus (BaYMV) was improved:The infected barley leaves were homogenized with high molarity (0.5M) potassium phosphate buffer pH7.0 containing 0.1% ME and 0.01M EDTA.After clarification with 1/4 V carbon tetrachloride, the virus was concentrated by precipitation with 6% PEG and 3% NaCl added 1% Triton x-100, and resuspended with the same buffer containing 0.5M urea, 0.1% ME and 0.01M EDTA, followed by ultracentrifugation with a 20% suctose cushion added 0.3% Triton X-100, and futher purified by 10-40% sucrose density gradient centrifugation.The purified BaYMV thus obtained showed the following characteristics:A260/A280 ratio of 1.20, Max.260/Min.240 ratio of 1.04.The yield of purifed BaYMV was about 5.5-8.0mg/kg infected leaves.Not any contaminats from host materials were observed in the purified preparation under electron microscopy.