Identification of necrophagous fly species from 12 different cities in China using ISSR and SCAR markers

Objective:To identify necrophagous fly speies from different regions in China using inter simple sequenc repeat(ISSR) and sequence-characterized amplified region(SCAR) melocular markers and to analyze their gene difference and genetic relationship.Methods:Five carrion fly species were collected from 12 cities and regions in China,including Musca domestica(M.domestica), Lucilia sericata(L.sericata),Chrysomya megacephala(C.megacephala),Helicophagella melanura(H.melanura),Boethcherisca peregrina,and they were studied using ISSR and SCAR markers.Results:Eight ISSR primers were used for amplification of 121 samples.679 clear and stable bands were identified,of which 516 bands were polymorphic.Several species-specific ISSR fragment were cloned and sequenced as an initial effort to derive the SCAR markers.Using M.domestica SCAR specific primers,SCAR-PCR amplification was performed for 8 M.domestca population sample DNA from different regions in China as well as L sericata,C.megacephala, H.melanura and Lucillia cupirina.The result showed only M.domestica produced specificalty 600 bp fragment,but L sericata,C.megacephala,H.melanura and Lucillia cupirina did not produce the same specific fragment.Clustering analysis showed clustering of most flies of M. domestica,C.megacephala and L sericata.M.domestica samples from different regions in China yielded different banding patterns.Conclusions:Application of ISSR-PCR and SCAR markers to identify necrophagous fly species from 12 cities and regions in China is first reported.ISSR-PCR and SCAR markers provide a quick reliable molecular marker technique for the identification of different species of necrophagous fly.