马铃薯腐烂茎线虫的鉴定及分子检测-植物病理专业毕业论文.docxVIP

度为0．5uM(上游引物／下游引物)，探针浓度为0．5¨M，镁离子浓度为3mM，其他 度为0．5uM(上游引物／下游引物)，探针浓度为0．5¨M，镁离子浓度为3mM，其他 的组分对整个程序的影响不大只要足量即可；最佳反应程序为： 95℃3min；95℃ 10sec，60℃25sec，40个循环；探针能够检测的最低浓度为5．0×10一¨g质粒DNA； 通过测试的8个不同种群的线虫，表明探针对Df啦，zc办螂如s驴“c幻r序列未缺失群体 具有很强的特异性，能有效的检测马铃薯腐烂茎线虫。操作简单，整个检测过程实行 自动化控制，完全闭管，消除了PCR假阳性的污染，无须PCR后处理。 - 关键词：马铃薯腐烂茎线虫腐烂茎线虫培养形态鉴定序列分析实时荧光检测 J - ，’ Identification Identification and Molecular Detection of Potato Rot Nematodep砂如玎c^“s d缸f，髓c幻，=) Abstract Potato I沁t Nematode was a kind of transfered entoparasitic nematode、Ⅳmch af．fect plant roots．they ate缸19i without higher plants．Potato Rot Nematode was a kind of impoIrtant patllogellic nematode．The nematodes occured in Beijing，Tianjin，Shandong， Hebei，Henan，Jiaulgsu，Zhej iaJlg，Fuj ian，Liaoning，Gansu，Hainall，aJld other proVinces in China． Mo印hological and molecular idemification for six stem nematode populations coUeted行om Beijing，heibei，jiansu proVince，China were researched．The ITS regions of r【)NA were sequenced and aIlalyzed．A real-time quantitatiVe polymerase chain reaction method for diagnosis of Df纱，Pnc而“s沈s驴甜c^Dr was deVeloped in present study．Main results or conclusions of the research were as f色llows： 1．A f色w methods and conditions culturing potato stem nematode were compared in present study．The result indicated：It was the best method that cultured the nematode with carrot dish．The optimized condition was at 25℃、70％relatiVe humidity under darkness． 2．rDNA-ITS sequence analysis of an s锄ples：the amplification of the rDNA-ITS 诧gion from t11ree stem nematode populations of Beij ing city， Daxing of Beij ing city，Funing of Hebei proVince(code Des一5、 Des-6、Des一7)in China and D．坊印cf矗om NetherlaIlds yielded one single fragment of 942bp，whereas the PCR products of ITS from three stem nematode populations of Zhangbei of Hebei proVince，Jiangsu proVince(Code Des一1、Des-3、Des一4)in China and from stem nematode group of Korea was 1 1 3 Obp．The ITS section was made sure by the sequence of rDNA—ITS region 1jrom GenBank．There was a 1 88bp of ab