Differentiation of Asian rice gall midge, Orseolia oryzae (Wood-Mason), biotypes by sequence characterized amplified regions (SCARs)

We developed a polymerase chain reaction (PCR)based assay that distinguishe d five different biotypes of the Asian gall midge (Orseolia oryzae), a majo r insect pest of rice. A total of 400 random primers were screened using ra ndom amplified polymorphic DNAs (RAPDs). Five diagnostic PCR products were isolated, cloned, sequenced and converted to sequence characterized amplifi ed regions (SCARs). Primers specific to these SCARs were able to amplify sp ecific DNA fragments from genomic DNAs of five biotypes of gall midge in a multiplexed-PCR-based assay. The amplified DNA fragments were used as diagn ostic markers to identify different biotypes of gall midge. The SCAR primer s were also capable of differentiating the Asian from the African rice gall midge (Orseolia oryzivora) as well as detecting a variant of biotype 5 whi ch caused an outbreak in Kerala, India. Unlike the use of plant host differ entials and midge feeding behaviour for identifying biotypes, this assay is fast, reliable and unaffected by environmental factors.