Behura S K, Sahu S C, Rajamani S, Devi A, Mago R, Nair S, Mohan M
Central Rice Research Institute (CRRI), Cuttack-753 006, Orissa, India.
Insect Mol Biol. 1999 Aug;8(3):391-7. doi: 10.1046/j.1365-2583.1999.83126.x.
We developed a polymerase chain reaction (PCR)-based assay that distinguished five different biotypes of the Asian gall midge (Orseolia oryzae), a major insect pest of rice. A total of 400 random primers were screened using random amplified polymorphic DNAs (RAPDs). Five diagnostic PCR products were isolated, cloned, sequenced and converted to sequence characterized amplified regions (SCARs). Primers specific to these SCARs were able to amplify specific DNA fragments from genomic DNAs of five biotypes of gall midge in a multiplexed-PCR-based assay. The amplified DNA fragments were used as diagnostic markers to identify different biotypes of gall midge. The SCAR primers were also capable of differentiating the Asian from the African rice gall midge (Orseolia oryzivora) as well as detecting a variant of biotype 5 which caused an outbreak in Kerala, India. Unlike the use of plant host differentials and midge feeding behaviour for identifying biotypes, this assay is fast, reliable and unaffected by environmental factors.
我们开发了一种基于聚合酶链反应（PCR）的检测方法，可区分亚洲稻瘿蚊（Orseolia oryzae）的五种不同生物型，亚洲稻瘿蚊是水稻的一种主要害虫。使用随机扩增多态性DNA（RAPD）筛选了总共400种随机引物。分离、克隆、测序了五个诊断性PCR产物，并将其转化为序列特征性扩增区域（SCAR）。在基于多重PCR的检测方法中，这些SCAR特异性引物能够从五种生物型稻瘿蚊的基因组DNA中扩增出特定的DNA片段。扩增的DNA片段用作诊断标记，以鉴定稻瘿蚊的不同生物型。SCAR引物还能够区分亚洲稻瘿蚊和非洲稻瘿蚊（Orseolia oryzivora），并检测出在印度喀拉拉邦引发疫情的生物型5变种。与使用植物寄主鉴别和瘿蚊取食行为来鉴定生物型不同，该检测方法快速、可靠，且不受环境因素影响。
