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In order to develop a rapid, sensitive and specific qPCR assay for detection and quantification of Tomato yellow leaf curl virus (TYLCV), a pair of primers and TaqMan probe were designed according to the conserved sequence of known TYLCV isolates. Combining with MNP technique, a novel MNP-qPCR detection method was established and verified based on specificity, sensitivity and reproducibility tests. The results indicated that the Ct value of plotted standard curve showed good linear relationship(R2 =0.9994)with the log of copy number of template. The established method showed a high specificity for TYLCV detection without crossing reaction with Tomato severe leaf curl virus and Tomato yellow leaf curl Sadinia virus, and was 10-fold more sensitive than routine PCR. Both coefficients of variation were less than 2%, indicating a good reproducibility. We have provided a novel method for detection of TYLCV in plant samples rapidly and quantitatively. 相似文献
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应用MNP-RT-PCR方法检测黄瓜绿斑驳花叶病毒 总被引：4，自引：0，他引：4
A novel RT-PCR method integrated with Magnetic Nano Particles (MNP), MNP-RT-PCR, was set up for detection of Cucumber green mottle mosaic virus (CGMMV). After the virus particles in crude sap were concentrated by MNP, viral RNAs were released and were detected by RT-PCR. CGMMV could be detected in as less as 10 ng watermelon leaf materials. Compared with normal RT-PCR, the method decreased the inhibitors of plant material and steps for extracting RNA, and also increased the sensitivity of RT-PCR detection in less time. The method is simple and suitable for quick detection of plant virus in a large number of samples. 相似文献
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靶向测序基因型检测（GBTS）技术及其应用 总被引：2，自引：0，他引：2
借助于分子标记进行基因型检测的技术在生物遗传改良等领域发挥着重要的作用。国际跨国种业公司凭借其高通量、自动化、大规模的共享检测平台,基因型检测技术得到广泛应用。随着从3G时代的高成本固相芯片和随机测序式基因型检测（genotyping by sequencing,GBS）发展到成本低、对检测平台要求较低、基于靶向测序基因型检测（genotyping by target sequencing,GBTS）的液相芯片,基因型检测技术完成了向4G时代的转变。在本文中首先介绍了两项最新的GBTS技术（基于多重PCR的GenoPlexs和基于液相探针捕获的GenoBaits）及其原理。同时,发展了可以在单个扩增子内检测多个SNP,称之为多聚单核苷酸多态性（multiple single-nucleotide-polymorphism cluster,mSNP或multiple dispersed nucleotide polymorphism,MNP）的技术,极大地提高了目标位点（扩增子）内变异的检测效率。与GBS和固相芯片相比,GBTS技术具有平台广适性、标记灵活性、检测高效性、信息可加性、支撑便捷性和应用广谱性。同一款标记集（例如玉米40K mSNP）,可以获得3种不同的标记形式（40K mSNP、260K SNP和754K单倍型）;并可以根据应用场景的需求,通过控制测序深度获得多种不同的标记密度（1—40K mSNP）。GenoPlexs和GenoBaits 2种技术相结合,可广泛应用于生物进化、遗传图谱构建、基因定位克隆、标记性状关联检测（全基因组关联分析——GWAS和混合样本分析——BSA）、后裔鉴定、基因渐渗、基因累加、品种权保护、品种质量监测、转基因成分/基因编辑/伴生生物检测等领域。目前,已经在20余种主要农作物、蔬菜以及部分动物和微生物中开发了GBTS标记50余套,并已广泛应用于上述领域。最后,展望了与未来GBTS应用相关的几个问题,包括便携式、自动化、高通量、智能化检测平台;根据用户需求定制的可变密度、多功能分子检测;GBTS与其他技术（KASP、高密度芯片、BSA策略等）的整合;基于资源共享的开源育种等。这些将推动GBTS技术在动物、植物和微生物遗传改良等领域的广泛应用。 相似文献
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