Preliminary Mapping The Freezing Tolerance Gene And Development Of Linkage Markers To Wild Potato S.commersonii

Potatoes(Solanum tuberosum L.)can be used as food and vegetables,and they are nutritious and comprehensive.Their production is second only to rice,wheat and corn,ranking fourth,and their output is the highest in non-cereal crops.China has a vast territory,and potato planting is planted in both north and south of China.This has led to different cultivation methods in China,and it will withstand various climate tests.Potatoes are often exposed to low temperature frosts in production,especially in the winter crops of southern China.Most of the seedlings of potatoes are from December to February,and this period is the coldest and most vulnerable to low temperature frost.Chinese potato is usually affected by low temperature frost during this period,and large-scale potato plants are affected by freezing damage,it would cause huge losses to the potato industry.Enhance the understanding of the cold-resistant mechanism of potato,increase the research on the cold resistance of potato,help the utilization of cold-resistant resources,and help to achieve certain breeding goals,which plays a very important role in China’s potato industry.In this study,the low temperature sensitive S.verrucosum was used as the female parent,and the diploid potato wild species S.commersonii with excellent cold resistance was used as male parent to obtain F1 hybrid and then backcrossed with S.commersonii to obtain the BC1 segregating population.The following main results:1.BC1 population phenotype identification: The LT50 value of 176 population progeny was identified by LT50 value method,and the phenotype of 328 population progeny was identified by field natural frost method.It was found that the population was separating phenotype to cold resistance.In the LT50 value of the population material determined by the LT50 assay,the LT50 values of the parental CMM5 and8341 were-5.17 ℃ and-3.91 ℃ respectively,and 33 of the 176 materials were resistant to cold materials(-7 ℃~-5 ℃).There are 71 sensitive materials(-4 ℃~0 ℃)and 72 intermediate materials(-5 ℃ ~-4 ℃).In the phenotype of 328 population materials identified by the field natural frost method,the Vega et al.(1995) method was used to divide the potato damage into 7 grades of 0-6,and the parental CMM5 and 8341 grades were 0.43 and 4.54,respectively.There are 155 cold-resistant materials(grade 0-2),94 sensitive materials(grade 3-6),and 79 intermediate materials(grade 2-3).Correlation analysis was used to analyze the correlation between the three repeated LT50 values,the three-year field natural frost phenotype,and the LT50 value and the field natural frost phenotype.It was found that there was only a repetition between the LT50 of the three replicates.There was a significant correlation between the LT50 value and the natural frost phenotype in the field,and there was a significant correlation between the phenotypes of the natural frost in the field for 3 years.Therefore,in this experiment,the field natural frost method is more suitable for the evaluation of cold resistance of VCC group materials.2.Separation of acclimation and freezing tolerance: Based on the data of the natural frost phenotype of the field for 3 years,24 parts of sensitive materials and 70 parts of cold-resistant traits were identified.However,the natural frosty plants in the field must have been subjected to long-term cold acclimation,so these 70 materials have freezing tolerance,acclimation or both.70 batches of materials were directly treated with low temperature frost,and no acclimation process was carried out.Through 4-5 rounds of repetition,25 direct cold resistant materials and 24 acclimated cold resistant materials were identified.The phenotype of the remaining 21 materials is not certain.3.BSA+DNA-seq analysis of extreme materials: 25 freezing tolerance materials were built into resistant pools,24 sensitive materials were set up as susceptible pools,and 24 acclimated resistant materials were set up as acclimation pools for DNA-seq sequencing.The pool sequencing depth was 40 x,the sequencing depth of the two parents is 15 x,and all pools get at least 42 Gb of data,and the parents get at least17 Gb of data.Using bioinformatics analysis,the average Δ(SNP-index)and G’ value distribution maps of potato freezing tolerance and potato acclimation resistance were plotted,and the peaks were located on chromosomes 2 and 5.Since the peaks of the average Δ(SNP-index)and G’ values of potato freezing tolerance and potato acclimation resistance were highly coincident,subsequent experiments did not distinguish between freezing tolerance and acclimation resistance of progenies.It is predicted that the cold resistance site is located at the position of chromosome36.23Mb-45.14 Mb by the position and height of the peak.4.Screening polymorphism markers for segregating population: 200 pairs of molecular markers were designed in the predicted freezing tolerance site of chromosome 36.23Mb-45.14 Mb.The 200 pairs of markers were verified by resistant and susceptible parents and two resistant and susceptible progenies respectively,the results show that 5 polymorphism markers were screened.5.Development of linkage markers to potato freezing tolerance: The correlation analysis of 5 polymorphism markers with 102 offspring phenotype,the results show that all screened markers were not linked to potato freezing tolerance.The follow-up work will continue.