迪卡配套系猪RYR1的PCR分析及其利用的研究

试验从鸡东县永安猪场随机抽取迪卡配套系猪B系和E系42头，通过PCR扩增技术和酶切鉴定．采用克隆技术进行测序，来检测是否含有氟烷基因序列。经过以上技术检测这42头猪均没有氟烷基因。     

单抗酶联试剂盒与PCR方法快速检测沙门氏菌的比较

本文对单抗酶联试剂盒与PCR方法快速检测沙门氏菌进行了比较。将沙门氏菌与大肠杆菌以不同比例混合后 ,分别用ELISA和PCR法检测 ,在 1∶1 0 0的比例时 ,用这 2种方法均能检测出阳性样品 ;在 1∶1 0 0 0的比例时 ,用ELISA法检测为阴性 ,而用PCR法则能检测出。检测鲜牛奶、龙虾、虾仁、熟食制品等样品 ,2种方法的符合率达 97 6 %。对冻虾仁、人粪便样品的前增菌、选择性增菌、后增菌过程进行同步检测 ,在样品的前增菌液 ,直接ELISA法检测的OD值小于 0 5 ,而PCR法扩增能出现特异条带 ,经国标方法验证为阳性 ,证实PCR法的敏感性优于直接ELISA法 ,而在选择性增菌液和M 肉汤后增菌液中 ,2种方法均能检出阳性样品。     

从海南省杂草胜红蓟和假马鞭上检测到粉虱传双生病毒

 利用三抗体夹心ELISA(TAS-ELISA)及PCR检测的方法对采自海南的胜红蓟(Ageratum conyzoides)和假马鞭(Stachytarphetajamaicensis)的5个病样进行了检测,表明均为粉虱传双生病毒。PCR扩增产物克隆后进行序列测定,结果表明存在2类双生病毒,其中样品Hn2存在2类病毒的复合侵染。这是在海南首次报道存在有粉虱传双生病毒。     

Production and characterization of a monoclonal antibody specific to the M serotype of plum pox potyvirus

A monoclonal antibody to an Albanian isolate of plum pox potyvirus (PPV) was obtained (MAbAL), that specifically recognized strain M of this virus. The specificity of MAbAL, assessed by comparative ELISA on 130 PPV isolates of different geographical origin, 22 of which were also tested by comparative IC-PCR, gave consistent and highly reproducible results. MAbAL seems to be elicited by a stable surface determinant that makes it particularly suitable for successful use under a wide range of conditions. MAbAL is an useful addition to the panel of PPV-specific MAbs available to date.     

Spread of potato virus Y in potato cultivars (Solanum tuberosum L.) with different levels of sensitivity

The aim of this work was to correlate the appearance of the symptoms, multiplication and spread of virus after mechanical inoculation of potato (Solanum tuberosum L.) cultivars showing different levels of susceptibility and sensitivity to Potato virus Y^NTN (PVY^NTN). The potato cultivars used were the resistant cultivar Sante and susceptible cultivars Igor, Pentland squire and Désirée. The spread of the virus PVY^NTN in infected plants was monitored using different methods: DAS-ELISA, tissue printing, immuno-serological electron microscopy and real-time PCR. In all three susceptible cultivars, the virus was detected in the inoculated leaves 4–5 days after inoculation. From there virus spread rapidly, first into the stem, then more or less simultaneously to the upper leaves and roots. Real-time PCR was shown to be very sensitive and enabled viral RNA to be detected in non-inoculated leaves of susceptible cultivar Igor earlier than other methods. Therefore, for exact studies of plant–virus interaction, a combination of methods which detect viruses on the basis of their different properties (coat protein, morphology or RNA) should be used to monitor the spread of viruses.     

sikFBA4基因启动子的克隆及低温表达模式分析

天山雪莲生长于常年积雪覆盖高海拔处的高山区域,具有很强的极端低温生境适应能力,属于研究植物低温适应的良好模式植物。前期的研究表明,sikFBA4基因可以显著提高番茄的抗寒能力。为进一步研究sikFBA4基因的耐寒响应模式,以天山雪莲为材料,采用实时荧光定量PCR分析sikFBA4在低温胁迫下的表达模式;利用高效热不对称PCR法(Hi-Tail PCR)克隆sikFBA4启动子序列并进行生物信息学分析。为分析PsikFBA4序列克隆的完整性及转录表达特性,将PsikFBA4与GUS基因融合在烟草中进行瞬时表达。结果表明,sikFBA4在低温胁迫条件下的表达发生瞬时显著上调,并在1 h达到峰值,而后表达下调。通过启动子克隆分析,在PsikFBA4序列-1648 bp位置处具有冷响应元件LTRE;进一步的GUS染色和酶活力测定结果表明,低温能够显著提高GUS基因的表达活性,说明sikFBA4属于低温诱导型基因。SikFBA4基因启动子的克隆、序列分析以及表达分析,为进一步探究雪莲果糖-1,6-二磷酸醛缩酶(sikFBAs)基因的表达与调控机制奠定了基础。     

山羊痘病毒SS株5个ANK基因缺失的重组病毒构建

为研究锚定蛋白基因(Ankyrin,ANK)对山羊痘病毒的影响,试验采用融合PCR和Overlap PCR技术扩增山羊痘病毒SS株5个ANK基因(ANK010、ANK138、ANK 140、ANK141.2和ANK145)两端侧翼序列和绿色荧光蛋白(GFP)基因,并将其产物连接Trans1-T 1载体构建ANK缺失的转...     

转基因小麦的定性PCR筛选检测技术

为了建立转基因小麦的PCR检测方法标准,以B73、B72、B102三个被转入外源高分子量谷蛋白亚基(HMW-GS)基因的转基因小麦品系为材料,对目前国内外转基因小麦中通用的标记基因bar和uidA、NOS终止子和Ubiquitin启动子进行了定性PCR的筛选检测,在国内外首次找到了小麦中特有的内参照基因麦谷醇溶蛋白基因(GAG56D),设计并合成了麦谷醇溶蛋白基因(GAG56D)和Ubiquitin启动子的引物序列,并对PCR反应条件进行了优化,PCR产物经过琼脂糖凝胶电泳分析后,可以检测到预期大小的目的片段.同时对PCR产物用实时荧光定量PCR进行了确证实验,并得到预期的结果.定性PCR最低检测灵敏度为0.5%(w/w).建立的转基因小麦定性PCR筛选检测方法具有通用性.