Construction Of Grape Mutation System And Identification And Evaluation Of Mutagenic Germplasm

Cultivars with high quality and special characteristics play a crucial role in improving grape fruit yield and quality.Aiming at the problems of weak stress resistance of grapes in China,it is of great significance to accelerate the selection of high-quality varieties with disease resistance,cold resistance,excellent fruit quality,and good wine making properties.In this experiment,the fresh food variety ’ Shine Muscat’ and rootstock SA15 were used as experimental materials,and colchicine was used as mutagen to analyze the mutagenic effects of different colchicine concentrations and different mutagenic time in cotton soaking and soaking methods.The mutant grapevines were identified by flow cytometry ploidy analyzer,and the morphological,metabolic,and physiological changes of the mutant grapevines were compared with diploid vines as controls.The red-fleshed superior line ’Jianhong 1’ induced by rootstock SA15 was used as the experimental material,and SA15 and ’ Merlot ’ were used as the control,the growth and development habits,fruit and wine quality,cold resistance and disease resistance of red-fleshed superior line ’Jianhong 1’ were comprehensively evaluated,and the whole genome resequencing technology was used to analyze the differences in fruit color between red-fleshed ’Jianhong 1’ and green-fleshed SA15,and to explore the variation genes associated with grape fruit color and the molecular mechanism of fruit redness.In this experiment,the technical system of grape variety mutagenesis,mutant identification and evaluation was established.The main findings are as follows:1.Establishment of polyploid mutation system and identification and evaluation of mutagenic germplasm.(1)The results of colchicine mutagenic showed that different colchicine concentrations and treatment time had different degrees of damage to the grapevines,and increasing the mutagenic concentration and prolonging the mutagenic time led to an increase in the mortality rate of the grapevines.Different mutagenic methods had different mutagenic effects on grapevines,the suitable concentration of colchicine cotton soaking method was0.8%,and the treatment time was 7 d,which led to a mortality rate of 36 % and the mutation rate of 1.3 %.(2)The results of ploidy identification showed that colchicine had different mutagenic effects on different varieties,and a mixed ploidy of diploid and tetraploid of ’ Shine Muscat ’was obtained by flow cytometry ploidy analyzer identification.No variation was obtained in SA15.The suitable variety for mutagenesis in this experiment was ’ Shine Muscat ’.(3)The comparison of physiological characteristics between mixed and diploid ’ Shine Muscat ’ showed that the metabolism and stress resistance physiological indexes of ’Shine Muscat ’mixed ploidy grapevines were significantly changed.Chlorophyll content,soluble sugar content,photosynthetic rate,chlorophyll fluorescence coefficient and antioxidant enzyme activity of adult leaves of mixed ploidy were significantly higher than those of diploid plants,and the malondialdehyde content was significantly reduced.2.Identification and evaluation of red-fleshed superior line induced by ethyl methane sulfonate(EMS).(1)The red-fleshed superior line ’Jianhong 1’ germinated in early April and matured in late September,compared with the control variety,the germination period was earlier and the maturity period was later.Compared with the control botanical traits,the number of leaf cracks of ’Jianhong 1’ was three,the shape of mature leaves was nearly triangular,and the leaves had a strong degree of bubble-like protrusions,the transverse diameter and longitudinal diameter of seeds were higher than those of the control,and reached a significant level.(2)The flesh color of ’Jianhong 1’ is purple-red and brighter than SA15 and ’ Merlot ’.The fruit grain of ’Jianhong 1’ was small,the single fruit weight was only 0.5 g,the fruit grain shape was nearly circular,and it was evenly distributed on the ear.The Soluble solid content of ’Jianhong 1’ fruit was 19.73%.,total sugar content was 26.57%,total acid content was11.51 g / kg,and Vc content is 167.29 mg / 100 g.In different parts of ’Jianhong 1’ fruit,the contents of total phenols and total flavonoids were in the order of peel > seed > pulp,the content of total anthocyanins was in the order of peel > pulp > seed,and the content of each phenolic substance was better than that of the control variety.(3)The evaluation results of cold resistance showed that after different lowtemperature stress,the germination rate of ’Jianhong 1’ was higher than that of the control variety,and it could tolerate a low temperature of-20°C.The results of disease resistance evaluation showed that the disease index of ’Jianhong 1’ was lower than that of the control variety SA15 and higher than that of the control variety ’Merlot’.The experiments proved that’Jianhong 1’ is a variety with high resistance to downy mildew and medium anthracnoseresistant varieties.(4)The analysis results of physical and chemical indexes showed that ’Jianhong 1’wine is dark red and has mixed fruit flavor.The alcohol content was 12.5 degrees,the residual sugar content was 5.34 g / L,and the titratable acid content was 10.70 g / L.Compared with the control variety,the content of phenolic substances in the wine sample was significantly increased,which had a certain winemaking advantage.(5)The sequencing results of ’Jianhong 1’ and SA15 showed that the average sequencing depth in ’Jianhong 1’ and SA15 were 24 × and 22 ×,a total of 11,455,416 SNP loci and 1,907,594 In Dels loci were detected between the two materials and eight mutant genes were screened out(DFR、ANS、AN11、GST、3GT、AT、OMT、CAS),which might enhance the ability of the fruit to synthesize anthocyanins and be the main reason for the purple color of ’Jianhong 1’ fruit.