New paper on prawn embryos about prepGEM™ (Zygem)

High-throughputDNAextractionforPCR-basedgenotypingofsinglePenaeusmonodonembryosandnauplii

CSIROFoodFuturesNationalResearchFlagship,CSIROMarineandAtmosphericResearch,Cleveland,QLD4163,Australia^b
CSIROFoodFuturesNationalResearchFlagship,CSIROLivestockIndustries,QueenslandBiosciencesPrecinct,StLucia,QLD4067,Australia^a
a,JeffA.Cowley^b,StuartJ.Arnold^aMinRao
 

2010PublishedbyElsevierB.V.

【Abstract】:AsimpletechniqueisdescribedforisolatingDNAfromindividualembryosornaupliioftheBlackTigershrimp,Penaeusmonodon.TheprotocolusedacommercialprepGEM™insectDNAextractionkitincombinationwithwaterbathultrasonicdisruption.DNAqualityandyieldsweresufficientformultiplexPCRdiscriminationofuptosixDNAmicrosatellitesinasingletest.Theutilityofthemethodwasdemonstratedindefiningtheparentageof1279embryos(7hold)spawnedfrom21femalesand684newly-hatchedN1toN2naupliispawnedfrom12femaleseachinseminatedartificiallywiththespermatophoresoftwodifferentmales.Overall,parentageassignmentsbasedonthesixmicrosatelliteMarkerswerededucedtobe89.5±6.7%and93.0±4.4%effectiveusingDNAisolatedfromindividualembryosandnauplii,respectively.Thetechniqueissimple,cost-effectiveandshouldprovidearobustmeansofisolatingDNAfromindividualembryosandnaupliiofaquaticinvertebratessuitableforPCR-basedgenotyping.

【摘要】：本文描述了一种从斑节对虾、黑虎虾的胚胎或幼体个体提取DNA的简单方法，主要是用商业化的prepGEM™昆虫提取试剂盒，同时结合水浴超声破碎。DNA质量及产量足够用于一次反应中多达6对微卫星DNA的多重PCR检测。此方法以已经在1279胚胎（7小时生命）亲子鉴定的实验中证实，这些胚胎来自21个雌体及各与2个雄体经过人工受精后的12个雌体新孵出的684个N1-N2幼体。基于胚胎或幼体个体DNA进行的6对微卫星标记的亲系鉴定，推导为89.5±6.7%和93.0±4.4%。此技术简单高效，节约成本，为水生无脊椎动物胚胎和幼体PCR基因分型提供了极佳的的DNA提取方法。