QTL Mapping And Candidate Genes Research For Grape Downy Mildew Resistance

Grape downy mildew is a multi-cycle fungal disease that caused by grape Plasmopara[Plasmopara viticola?Berk.et Curtis?Berl.et de Toni.],as one of the most important fungal diseases in grape production,it has been distributed throughout the world in grapes areas,high temperature and humidity are mainly factors to make this it epidemic.In this experiment,we use two interspecific F₁ offsprings as plant materials,one of them was derived from Vitis vinifera'Red Globe'and Vitis amurensis'Shuangyou',another was derived from Vitis vinifera'Chardonnay'and‘Beibinghong',and‘Beibinghong'was derived from Vitis vinifer×Vitis amurensis,then we conducted the Grape downy mildew resistance identification between 2 F1 offsprings,and completed QTL of grape downy mildew resistance traits.With the analysis of candidate genes from the major QTL interval,candidate resistance genes and the expression level of those genes verified by fluorescence quantitative PCR.The main results were shown as follows:We identified the resistance of the grape downy mildew in two F1 generations leaves in two consecutive years indoor,one of them was derived from Vitis vinifera'Red Globe'and Vitis amurensis'Shuangyou',which have 149 strains,another awas derived from Vitis vinifera'Chardonnay'and‘Beibinghong',which have 130 strains.The results showed that:In 2016 and2017,'Red Globe'and'Chardonnay'showed high susceptibility,average disease index was85.56 and 70.37;‘Shuangyou'and‘Beibinghong'showed high resistance,the average disease index was 1.48 and 0.56 respectively.The offsprings showed five levels of immunity,high resistance,resistance,susceptibility,and high susceptibility.With multiple environments in two years show that,the average phenotypes of grape downy mildew displayed as 2.03%,27.70%,48.31%,17.57%,4.39%at F1 offsprings by‘Red Globe'בShuangyou',of which78.04%and 21.96%were higher than resistance level and susceptible level,and 3 strains showed immunity;In addition,the average phenotypes of grape downy mildew resistance displayed as 0,3.08%,48.85%,40.39%,7.69%in F1 offsprings by'Chardonnay'בBeibinghong',of which 51.93%and 48.07%were higher than resistance level and susceptible level,and didn't have immunity strains in F1 generations.On the whole,the resistance of F1 offsprings of grape downy mildew showed that the‘Red Globe'בShuangyou'was exceeded than‘Chardonnay'בBeibinghong',resistance heredity of offspring affected by their parents.Additionally,according to the phenotypic distribution ratios of the individual plants in the two families,it can be seen that the disease resistance is significantly separated,showing a continuous distribution,showing the character of quantitative traits,and the resistance evaluation of phenotypic value can be used for downy mildew QTL mapping.The genetic map was constructed by‘Red Globe'בShuangyou'and‘Chardonnay'בBeibinghong'F1 offsprings.Grape downy mildew resistance QTL mapping were then achieved by using MapQTL5.0 software and combined with downy mildew resistant phenotype identification of two years,choose the interval mapping method to determine the standard LOD?3 as a QTL site.The results showed that 26 locations associated with the resistance of downy mildew.We found 7 QTL at'Red Globe'בShuangyou'integration genetic map,the result of 2016 were mainly located at LG15,and the result of 2017 were mainly located at LG11 and LG15.In addition,19 QTLs were found at‘Chardonnay'בBeibinghong'integration genetic map in 2016 and 2017 and located at LG1?LG5?LG7?LG9?LG13?LG17.According to the downy mildew resistance QTL mapping results,five candidate genes were selected including MAPK,Vv-LRR,TIR,RPM1 and F-box in‘Red Globe'בShuangyou'and‘Chardonnay'בBeibinghong'F1 offsprings,then,primer design was performed for these five candidate genes and obtained the final target fragment sizes:159 bp,215 bp,224 bp,225 bp and 174 bp.Finally,the application of real-time fluorescence quantitative PCR to analyze the expression of genes in disease-resistant and susceptible grape leaves,the result showed that these candidate genes were induced by downy mildew and the expression level in disease-resistant varieties was higher than susceptible varieties.These candidate genes may be potential grape downy mildew genes,and the result will laying the theoretical foundation for molecular marker-assisted selective breeding of grapevine.