摘要:
应用液氮冷冻香蕉束顶病组织,高速粉碎,氯仿—正丁醇澄清,差速离心,沉淀悬浮后经蔗糖垫部分纯化病毒,收集的蔗糖垫浓缩后经0—40%连续蔗糖梯度离心4小时,在离心管上半部形成一个明显的含有大量等径病毒的紫外吸收峰。再经不连续硫酸铯梯度离心6小时,形成一狭窄的乳白色带,含有高度纯化的直径约18nm的香蕉束顶病毒(BBTV)。用台湾提供的BBTV单克隆抗体,ELISA法可检测到提纯过程中病毒的存在,ISEM法可从粗提物中捕获大量等径病毒。ELISA法也用于检测田间及组织培养原种苗的带毒情况。
Abstract:
Banana bunchy-top virus (BBTV) has been purified from diseased banana tissues in Fujian province,China.The banana tissues are pulverized with liquid nitrogen in a coffee grinder.The extracts are clarified with chloroform-butanol,and concentrated by one cycle of differential centrifugation.The suspension of the pellets is further centrifuged on the top of two layer sucrose cushions with 10% and 30% concentrations.The particles in the cushions are collected, concentrated, resuspended and centrifuged on the top of 0 - 40 % sucrose density gradient.The virus particles in a UV-absorbing band on the uppper tube of the sucrose density gradient are further purified in a discontinuous density gradient of cesium sulfate and form a narrow visible band containing a large number of highly purified isometric virus with the diameter about 18 nm.The virus is able to be detected in the process of purification by ELISA with the monoclonal antibody a-gainst BBTV which provided from Taiwan University, and be trapped by ISEM from the clarified extract.The monoclonal antibody has been used to diagnose the field-growing banana samples and tissue culture seedlings.
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