【摘要】 将小麦黄色花叶病毒(wheat yellow mosaic virus,WVMV)外壳蛋白基因克隆到pET-30a(+)上,获得E coil表达载体pETWCP。SDS-PAGE和Western blotting分析结果表明,pETWCP在E coil中经IPTG诱导后,可特异地表达38.5 kD的融合蛋白,光密度扫描分析结果,融合蛋白表达量约占细菌总蛋白的40％。以诱导表达产物作为抗原,免疫家兔制备了特异性强的抗血清,用微量沉淀法测其效价为1:1 024。该抗血清可替代常规病毒粒子抗血清,用于酶联免疫吸附测定、免疫电镜和Western blotting检测WYMV。更多还原

【Abstract】 The prokaryotic expression vector pETWCP containing wheat yellow mosaic virus (WYMV) coat protein (CP) gene was constructed by cloning the CP gene into pET-30a(+) and transferred into E. coli BL21(DE3) pLysS. The results of SDS-PAGE and Western blotting analysis showed that specific expression of a 38.5 kD fusion protein was achieved by IPTG induction. The expression product was accumulated up to 40% of the total cellular protein estimated by densitometer gel scanner. The fusion protein was purified from SDS-polyacrylamide gels and antiserum against the protein was raised in rabbit.The titer of antiserum was 1:1 024 estimated by miniprecipitatioa The antiserum was used to substitute for regular antibody against virion, for specific detection of WYMV in plant extracts by ELISA, ISEM and Western blotting analysis respectively.更多还原