| 土豆是仅次于水稻、小麦、玉米的世界第四年夜食粮作物。我国事土豆临盆的年夜国，栽种面积与产量均居世界第一，但我国的土豆家当根本还处于一种自觉的无序状况，与国外种薯质量差别伟大，各类土豆病害产生情形严重。是以从土豆临盆强国引种高品德土豆成为晋升我国土豆家当的主要门路和手腕。根据FAO和IPPC的相干划定，法语论文网站，经由过程引种前的无害生物风险剖析，可以迷信有用的下降引种带来的各类风险和伤害，掩护本国农业临盆、生态平安。今朝在土豆上已发明的病毒、类病毒多达30余种，病毒病的产生和流传严重制约了土豆的临盆。树立疾速、精确、敏锐的病毒检测办法可以最年夜水平切实其实保引种土豆的种薯质量，掌握土豆病毒病的流传。本研究经由过程无害生物风险剖析对从法国、荷兰、俄罗斯引种土豆种薯停止了公道的风险猜测，并确立了迷信有用的风险治理办法，下降了引种风险；同时树立了土豆病毒的单重、两重、多重反转录聚合酶链式反响检测系统。依据FAO制订的无害生物风险剖析准则，从流传门路开端停止无害生物风险剖析，法语论文题目，初步提出了法国、荷兰、俄罗斯三国进境土豆种薯上的无害生物品种和数目，其分离为179种、152种、142种。依据无害生物的地舆散布、官方掌握、经济主要性、种薯传带的能够性等综合目标停止初步评价，挑选出进境土豆种薯潜伏的限制无害生物（PRP）和非限制无害生物（NRP）。对进境土豆种薯潜伏的PRP，依据无害生物传入、定殖、分散的能够性和潜伏的作用等停止具体的风险评价，肯定了列国进境土豆上潜伏的检疫性无害生物（QP）和限制的非检疫性无害生物（RNQP）。剖析肯定法国QP共14种、RNQP共23种；荷兰QP共26种、RNQP共26种；俄罗斯QP共21种、RNQP共17种。根据风险剖析成果，拔取3种有较高引种风险的土豆病毒一土豆M病毒（PVM）、番茄斑萎病毒（TSWV）、土豆纺锤块茎类病毒（PSTVd）停止RT一PCR检测。运用试剂盒法、RNAiso Plus试剂法提取土豆病毒RNA，成果注解二者提取的RNA纯度高且完全性较好，知足RT一PCR检测的请求。依据NCBI报导的病毒全基因组序的守旧区域序列设计了PVM、TSWV、PSTVd特异性引物对，采取单重RT一PCR对3种土豆病毒停止了扩增，扩增片段年夜小分离为630bp（PVM）、747bp（TSWV）、251bp（PSTVd）。收受接管3种病毒的单重扩减产物并测序，经NCBI BIAST同已报导的病毒序列比对，证实PVM、TSWV、PSTVd的RT一PCR扩增靶带序列完整准确。实验从反转录下流引物量、Mg2/浓度等方面临单重、两重和多重RT一PCR系统检测土豆病毒停止了摸索和优化。 Abstract: Potatoes are after rice, wheat and Maize in the world's fourth largest food crop. I state potato production big country, planting area and production ranks first in the world, but China's potato belongings haven in a conscious of the disordered state, and foreign seed potato quality difference is great and all kinds of potato diseases produce situation is serious. Is the main way to be introduced from the wrist and potato production promotion in China power high quality potatoes have potatoes. According to FAO and IPPC relevant provision, through the introduction of harmless biological risk analysis, scientific and effective decline introduction to bring all kinds of risks and damage, cover their agricultural production and ecological security. Today the potatoes have been invented of virus and up to more than 30 kinds of virus disease of generation and spread seriously restrict the birth of potatoes. To establish a rapid, accurate and sensitive detection method can be the most effective way to effectively protect the quality of potato chips, potato virus spread. In this study, through the process of biologically sound risk analysis of from France, the Netherlands, Russia introduced potato seed stop the reasonable risk speculation, and establish a scientific and effective risk control measures, decreased the introduction risk; and set up a potato virus of single, double and multiple reverse transcription polymerase chain reaction detection system. Basis for formulating FAO of the harmless biological risk analysis principles, from the spread of roads beginning of harmless pest risk analysis, initially proposed the France, the Netherlands, Russia imported potato seed tuber of harmless biological variety and number. The separation is 179, 152 and 142 species. According to the harmless bio geographical spread, the official master, economy, seed transmission belt to comprehensive target of the preliminary evaluation, selected imported potatoes sweet potato latent restrict harmless biological (PRP) and non harmless organisms (NRP) limit. The imported potatoes sweet potato latent PrP, harmless biological basis for afferent and colonization, dispersion can and latent influence to stop a specific risk assessment, affirmed the nations with the entry of potato latent quarantine pests (QP) and limit non quarantine harmless biological RNQP. Analysis of 14 kinds of QP in France, there are 23 kinds of RNQP, Holland QP a total of 26, RNQP a total of 26, 21 kinds of Russian QP, RNQP a total of 17 kinds of. According to the risk analysis of the results, select the three have a higher risk of introduction of potato virus a potato virus M (PVM), tomato spotted wilt virus (TSWV) and potato spindle tuber virus (PSTVd) stop RT - PCR. Application kit method, Plus RNAiso reagent extraction of potato virus RNA, the results of the extraction of the two extracted RNA high purity and complete better, meet PCR a RT test request. According to the sequences of the reported in NCBI virus genome sequence of conservative region design the PVM, TSWV, PSTVd specific primers to take single RT - PCR of 3 kinds of potato virus to stop the amplification and separation of small large fragments amplified for 630 (PVM), 747bp (TSWV) and 251bp (PSTVd). The single re expansion and sequencing of the 3 viruses were received and sequenced. The sequences of, TSWV, RT and PCR amplified by BIAST were confirmed to have a complete and accurate sequence of PVM, PSTVd and NCBI. Experiments from the reverse transcription of the low primer, Mg2/ concentration and so on are faced with a single, double and multiple PCR a RT system for detection of potato virus to stop the exploration and optimization. 目录: |
