Research On RAPD Markers Linked To Powdery Mildew And Downy Mildew Resistant Gene In Chinese Wild Vitis Pseudoreticulata

F₁ and F₂ populations from the cross between resistant Vitis pseudoreticulata W.T.Wang clone Baihe-35-1 and susceptible Carignane(V.vinifera L.) were observed for downy mildew under a natural setting condition.The inheritance of resistance in Chinese wild Vitis to downy mildew was analysed.On this basis,the screening for new RAPD markers correlated with the resistance to downy mildew in Chinese wild Vitis were carried out.Additionally,through the sequence analysis of the obtained markers,the RAPD markers were proved to be useful in marker assisted breeding,moreover,can be used in construting genetic maps,and the cloning of the resistant genes utilizing the genetic maps hereafter.Besides,the validation of RAPD markers linked to resistance to powdery mildew was processed based on the enlarged population size.The results are as follows:1.A total of 520 random sequence decamer oligonucleotide primers were used to screen for RAPD markers linked to downy mildew resistance in Chinese wild Vitis.Six RAPD markers were obtained in donor parent Baihe-35-1,and then cloned and sequenced.The length of the fragments are 369 bp,527 bp,615 bp,677 bp,1224 bp and 725 bp,respectively. They were designated as S294-369,S202-527,S382-615,S221-677,S416-1224 and S390-725,respectively.2.Co-segregation analysis of these 6 markers in F₂ progenies confirmed that all markers were linked to genes which control downy mildew disease resistance.The coincidence rate of the markers to phenotype are 72%,68%,75%,71%,69%and 71%,which therefore provided a molecular basis to marker assisted selection in grape disease-resistant breeding.Linkage analysis of the six markers via Joinmap 3.0 with LOD=3.0 indicated that,the six markers have no linkage relationship between each other,and therefore,can be targeted in different linkage groups.For this we can putatively say that these six RAPD markers respectively marks different downy mildew resistant genes which are located in different linkage groups.3.Annotation of the six marker fragments was performed,and the results indicated that the six RAPDs are all highly homologous,between 94%-98%,to the whole genome shotgun sequences which have been landed into GenBank recently.The BLASTx results showed that S382-615,S416-1224,S390-725 all encode unnamed grapevine proteins,among which,the peptides translated from sequence S382-615 have 56(56/86,65%) AA similar with transposase-like protein in Musa acuminata,51(51/88,57%) AA similar with hAT(histone acetyltransferase) dimerisation domain-containing protein in Arabidopsis thaliana.The S390-725 translated nucleotide has 31(31/46,67%) AA with(The haloacid dehalogenase, HAD) subfamily IB hydrolase hyperthetical I.The function of this enzyme is currently unknown,nevertheless,it was thought putatively to be correlated with antibiotic tolerance in bacteria and disease resistance,adversity tolerance in plant.Sequences of the six fragments were landed into GenBank,with the IDs:FI107280,FI107282,FI107279,FI107281, FI107277 and FI107278,respectively.