Eachkitcontains20individual10-merprimers(suppliedataminimumamountof50nmoleperprimer).TheseprimersaresuitableforuseingeneticmappingandDNAfingerprinting.IntheRAPDtechnique,asingle10merofarbitrarysequenceisusedasaprimerinPCRtoamplifygenomicDNAwherethesequenceoftheDNAiscompletelyunknown.GenomicDNAfromdifferentindividualsgivesdifferentPCRproducts,allowingtheidentificationofDNApolymorphismsthatcanbeusedtoidentifydifferentindividualsorasgeneticMarkers.|ThenumberofPCRproductsgeneratedfromeachgenomicDNAsampledependsontheprimersequence,thegenomicDNAsequence,andthegenomesize.Assumingthattheprimingsitesofa10merRAPDprimerarerandomlydistributedthroughoutagenome,thetheoreticalnumberofPCRproductsisapproximately2.5x10e9xG,whereGisthesizeofthehaploidgenomeinbasepairs.Usingthiscalculation,ahaploidgenomeof2x10e9bp,forexample,isexpectedtogiveabout5PCRproducts.Thispredictionisincloseagreementwithexperimentalresults.
KitComponents
t01 CACACCGTGTtt11 ACCGTGCCGT02 TGACCAGGCAtt12 GTCCTCGTGT03 ACGGTTCCACtt13 CACGGACCGA04 GTCTTGGGCAtt14 TCGCAGCGTT05 GTCACCTGCTtt15 CTGCAATGGG06 GGCGCGTTAGtt16 AACCCTTCCC07 GACGAGCAGGtt17 AGTTCCGCGA08 GGCTGCCAGTtt18 GTTGCGCAGT09 TGGAGTCCCCtt19 TGACAGCCCC10 CCCGTCTACCtt20 TCTGCCTGGA
Storeat-20°C.Lyophilizedpowderisstablefor6monthsat-20°C.Formaximumrecoveryofproduct,centrifugetheoriginalvialafterthawingandpriortoremovingthecap.
References
1.Grundmann,Towner,Dijkshoorn,Gerner-Smidt,Maher,Seifert,Vaneechoutte.MulticenterstudyusingstandardizedprotocolsandreagentsforevaluationofreproducibilityafPCR-basedfingerprintingofAcinetobacterspp.J.Clin.MicroBIOLOGy35(12):3071-3077(1997).2.Berg,D.,etal.,MethodsinMolecularandCellularBiology5:13-24(1994).3.Welsh,J.,McClelland,M.,FingerprintinggenomesusingPCRwitharbitraryprimers.NucleicAcidsRes.18(24):7213-7218(1990).4.Williams,J.G.,Kubelik,A.R.,Livak,K.J.,Rafalski,J.A.,Tingey,S.V.,DNApolymorphismsamplifiedbyarbitraryprimersareusefulasgeneticmarkers.NucleicAcidsResearch18:6531-6535(1990).5.Micheli,M.,etal.(1997).InFingerprintingMethodsBasedonArbitrarilyPrimedPCR,pp.55-63.Springer-VerlagPress,NY.6.Graves,L.,Swaminathan,B.(1993).InDiagnosticMolecularMicrobiologyPrinciplesandApplications,pp.617-621.ASMPress,Washington,DC.
