Molecular Markers Of Resistance Genes To Tomato Gray Leaf Spot And Verticillium Wilt In Tomato And Screening For Germplasm Resource

Tomato gray leaf spot and Verticillium wilt are two major diseases that often occur in tomato production.The Sm and Ve genes are the major disease resistance genes of tomato gray leaf spot and Verticillium wilt,respectively.In this study,resistant homozygous(Sm/Sm)?resistant heterozygotes(Sm/sm)and susceptible homozygous(sm/sm)were used as experimental materials,based on the published RFLP markers tightly linked to Sm,designed specific primers for PCR amplification.Sequence analysis of PCR amplification products were performed to find the restriction sites.A new CAPS marker named “T050” was created which was closely linked to the disease resistance gene Sm of tomato grey leaf spot.Resistant homozygous(Ve/Ve)? resistant heterozygotes(Ve/ve)and susceptible homozygous(ve/ve)to tomato verticillium wilt were used as experimental materials,based on the published literature,designed specific primers for PCR amplification.We created "Ve-1" SCAR marker,which is tightly linked to resistance gene Ve.On this basis,the validity of the two molecular markers was verified.The "T050" and "Ve-1" markers were applied to the selection and identification of tomato breeding materialsCAPS marker "T050" : The experimental materials were amplified by PCR using specific primers T050F/T050 R,and all the experimental materials were amplified with the same specific fragment of 651 bp.After digested with Scr F I enzyme,the resistant homozygous materials showed a fragment of 651 bp,resistant heterozygotes materials showed three fragments of 651 bp,355 bp and 296 bp and susceptible homozygous materials showed two fragments of 355 bp and 296 bp.Primer sequences are T050F: 5'-ATC ACC CTT CTG CCT TTT CTT C-3';T050R: 5'-ATC TGG GAT CTG AAT GTC ATC CTC-3'.The marker "T050" as verified by using 9 different genetic backgrounds and tomato varieties from different sources as experimental materials.The verification results indicated that the marker is a clear and stable codominant molecular marker.A total of 91 unknown genotypes of tomato materials were selected by marker "T050".As a result,20 resistant homozygous materials and 6 resistant heterozygotes materials were screened.SCAR marker "Ve-1" : The experimental materials were amplified by PCR using specific primers T050F/T050 R,the resistant homozygous materials showed a fragment of 1050 bp,resistant heterozygotes materials showed two fragments of 1050 bp and 1000 bp and susceptible homozygous materials showed a fragments of 1000 bp.Primer sequences are Ve-1F: 5'-CCA TGA ACA GAT GTG ACT TGT GTG-3';Ve-1R: 5'-CGA TAA CAA CCA ACT CTC TGT CTC-3'.The marker "Ve-1" as verified by using 12 different genetic backgrounds and tomato varieties from different sources as experimental materials.A total of 120 unknown genotypes of tomato materials were selected by marker "Ve-1".As a result,94 resistant homozygous materials and 3 resistant heterozygotes materials were obtained.The molecular markers of disease-resistant gene "T050" and "Ve-1" obtained in this study provide technical support for tomato disease resistance breeding.Disease-resistant germplasm obtained lays a foundation for further breeding new tomato varieties with complex disease resistance.