Preparation And Application Of Polyclonal Antibodies For Three Plant Viruses Using Prokaryotic Expressed Viral Coat Protein

The development of modem plant virology has gone through hundreds of years,the new era of agricultural production requirements are getting higher and higher,but the situation tends to be complicated.This requires plant virus detection methods and methods of operation to be more and more fast,more sensitive and accurate,but also more and more simple and more innovative.Plant virus detection in the entire plant virus research occupies a very important position,serological detection is one of the ways.A polyclonal antibody was prepared for the virus which was more prevalent and had a greater risk for the detection and identification of the virus.Tomato chlorosis virus(ToCV)belongs to the genus of Clinoviridae,which is the genus in the family of Crinivirus.The genome is a bimodal linear mononuclear RNA,and the coat protein gene is located on RNA2.It has a wide variety of host types and covers a wide range of worldwide epidemics,causing serious economic losses.Because of its extensive spread of the vector,as well as the initial symptoms of diseased plants is not obvious,resulting in its difficult to control.At present,the method of identifying ToCV is roughly visual method,RT-PCR detection method,reverse transcription-mediated isothermal amplification technique(RT-LAMP)detection method,tissue blot hybridization and serological detection method.In this study,we used the method of RT-PCR to link the amplified virus shell sequence with the vector to construct the tomato protein gene expression vector of chlorophyll virus coat protein.The recombinant protein was expressed as prokaryotic antigen Immunization of New Zealand male rabbits was used to prepare the antiserum.Serum was collected by indirect ELISA.Western blot was used to determine its specificity.The titer of the antiserum is about 1:50 and has certain specificity.The detection and identification of ToCV has practical application value.Bean common mosaic virus(BCMV)is a virus of Potyvirus,all over the world.BCMV gene structure is typical of the potato Y virus gene structure,the virus genome has only one strand of RNA.Under natural conditions,BCMV infects many plants,in which the beans are more severely affected.The virus has a variety of ways to spread,both through aphids for non-persistent way of poisoning,but also through the seeds and pollen for poisoning.The source of this study was derived from JiNing,Shandong Province,and the expression vector of the expression gene was constructed.The same method was used to express the purified protein to obtain the serum titer.The titer of the serum was determined to be 1:1000,and the results of Western blot showed that it was specific and could be identified by detection of left and right BCMV.Capsicum chlorosis virus(CaCV)is a tentative species of the genus Tospovirus and belongs to the watermelon silver mottle virus(WSMoV)serum group.First,Australia reported the discovery of the virus,followed by Thailand,India,successive reports found that the infection occurred until a wide range of diseases found outbreaks.The biological material used in this study was collected from Guangdong Province,and the prokaryotic expression vector of the virus shell was constructed by RT-PCR.The prokaryotic expression and the preparation of polyclonal antibody were carried out.The serum titers of the two rabbits were determined to be 1:10000,1:20000,and they were specific and could be used in the actual virus detection and identification.