Prokaryotic Expression Of CP Gene Of Sugarcane Mosaic Virus Strain E And Preparation Of Antibody

Sugarcane is the most important sugar crop and is accounting for more than ninety percent of sugar products in our country. At the same time, sugarcane is the best-regenerated energy crop for production of fuel ethanol in south China. Sugarcane mosaic virus (ScMV) is one of the most serious sugarcane diseases in China and in the world. It causes severely decline in quality and quantity of sugarcane and is responded to variety retrogression. Breeding and cultivation of disease resistant varieties are the most effective measures. Cultivation of disease free plants is in the next and will result in more than thirty percent increment of sugarcane stalk product. In spite of the most effective method for breeding of disease resistant varieties in sugarcane is the usage of traditional sexual crossing; variety improvement by means of gene engineering has been proven as an effect.Supported by Grant from China High Technology (863) Project (2002AA241031) and National 948 Project (2003-Q06), sugarcane transformation of gene responded to coat protein (CP) of ScMV strain E (ScMV- E) has been carried out in Key Lab of Eco-physiology and Genetic Improvement for Sugarcane of Ministry of Agriculture. Some transgenic sugarcane plants are gained and middle test in field is carried out by permission. High specific antibody responded to ScMV-E CP is the necessary for research on the expression of specific protein responded to ScMV-E CP gene. In the other side, the method of ScMV detection with quickly and sensitivity is necessary for the production and application of ScMV free sugarcane plants. The technology of antibody immunity is the best method. Construction of the prokaryotic expression vector contained the target gene of ScMV-E CP gene is carried out and expressed in Escherichia coli BL21 (DE3). The purified CP of ScMV-E extracted from ï¿¡ coli BL21 (DE3) acts as immuneogen to prepare polyclonal antibody. Main results are as follows.(1) The recombinant plasmid pNUSCP with target CP gene of ScMV-E was amplified by specific primers with the sites of double restriction enzyme digestion, BamH I and Sal I, respectively according to CP gene sequence of ScMV-E and the polyclonal sites of prokaryotic expression vectors, the target CP gene fragment of ScMV-E was obtained by PCR amplification and was...