Studies On The Tissue Culture System Establishment And Elimination And Detection Leafroll Virus Of Four Grapes

Grapevine leafroll disease is the most devastating disease of grapevine in many grapeving producting areas. it distributes widely and creates to the biggest economic loss, besides, it is the only one that has a nature transmission vector. It have the vital practical significance in improving production of grape to take the treatment of virus elimination and promotes the non-viruses cultivation.Four represent varieties The Beauty, Excellent Seedless, Red Earth, Ruby Seedless with virus were used as experiment materials and they are kinds of Hunan commonly cultivated Vitis Vinifera L grapes. Domestic or international scholars had not studied on these 4 varieties with virus elimination. Biology or serology method are usually used to detect their results about the rate of other grape varieties virus elimination. Because of the lower virus density in the plants vivo, detection results appears the false negative phenomenon, draws the wrong viral eliminating conclusion, therefore this experiment uses the RT-PCR detection technique to detect Plantlets of Virus Elimination. Therefore, studying on the methods of elimination and RT-PCR detection techniques can provide reliable guidance for production of application.The results as follows:1. The tissue Culture system had been Setted up for Ruby Seedless, Red earth, Beauty Mean, Excellent Seedless. The base or the middle of the stem was more stiutable for bud sprouting and about September 10 was the best suitable implanting time. The fittest optimum medium for the stem's initial culture,multiplication and for rooting have been founded for the four grape varieties. Domesticating Rooting Plantlet in sealed tube 6to 8 days and then Domesticating in unsealed tube 10days can improve the survival rate of transplanting. It is an basic for studying on techniques of elinination by providing plently tissue culture planlets and obtaining virus-free planlets.2. The shoot tip culture, Shoot-tip Implanting after thermotherapy and 4℃Culture,Shoot-tip side grafting, medication therapy are adopted to study the four methods of elimination. Consider the two factors of survival rate and the rate of elimination synthetically, the survival and eliminated shoots rate of thermotherapy combine with shoot tip culture has raise obviously than single shoot tip culture,besides, The ratio of eliminated shoots achieved by the method of Constant Temperature is better than Thermotherapy Alternating Temperature Thermotherapy. 4℃Culture for a long time can eleminate virus ,and it is simple and easy to do to operate, so it can be regarded as the effectual new method to eleminate virus of the plant. Shoot-tip side grafting plants also can eleminate GlRav virus, the resisting is strengthened after grafting and has obviously accelerated the growth speed of nutrition after transplanting. Medication therapy has good passivation function on the virus, but there is certain influence to Shoot-tip in the progress of growthing.3.Using the leaves of non-virus planlets as materials and microanalysis method for RNA extraction . According sequence reported GLRaV gene to design and formate the guide thing, regard RNA drawn as the template, formate cDNA and regard cDNA as the template to amplify PCR, the peculiar passage which gets GLRaV is 300bp, a large number of samples with virus can be measured at the same time.