Molecular Markers And Mapping Of Low Temperature Sensitive Genes In Wheat Leaves

Low temperature is one of the main factors that cause crops to reduce production.In northern China,the temperature in winter is low and the intensity of the cold current is large.Wheat frost damage is often caused,and yield is also greatly affected.Low temperature frost damage ha s also become the biggest natural disaster in wheat production.Common wheat varieties show resistance to low temperatures above 0 degrees.Leaf low temperature sensitive mutant BN044371 is a natural mutant found during wheat breeding.It is sensitive to temperatures above 0?and around4?.The leaves appear yellow spots,but as the temperature rises and the plant develops,the leaf color returns to normal.This trait is called Wheat Leaf Chilling Sensitivity.The leaves of this m utant plant are sensitive to low temperature and the symptoms of leaf color change.At present,th ere are many cold resistance genes found in wheat,but due to the insufficient number of genes di scovered and the slow research of signal transduction mechanisms related to low temperature,people have very limited understanding of their resistance mechanisms.Therefore,only by digging more genes related to low temperature and studying their functions,it is possible to lay the foundatio n for studying the mechanism of low temperature resistance of wheat and the cultivation of new wheat varieties.Therefore,this study has important research and utilization value to analyze and lo cate the genetic law of low temperature sensitive genes in wheat leaves.In this study,the natural mutant BN044371 was taken as the research object,and the genetic law of low temperature sensitive genes was analyzed.They were crossed with the low-tempe rature sensitive parents Bainong4199 and Bainongaikang58 to construct F₁,F₂ segregated populations and F_2:3:3 families,and the genetic rules of low-temperature sensitive genes?temporarily named wc hs1 gene?in wheat leaves were clarified.The F₂ segregated population of the low-temperature sensitive parent BN044371 and the low-temperature sensitive parent Bainong4199 were used as experimental materials,Using next-generation sequencing technology to establish low-cost and high-efficie ncy mixed pool transcriptome sequencing?Bulked segregant RNA-Seq,BSR-Seq?technology for wheat,combined with the self-developed enzyme-amplified polymorphism sequence molecular marker?CAPS?,the molecular markers of the wchs1 gene were initially located.The results were listed as following:1.Performance of leaf low temperature sensitive phenotype traits in fieldAfter sowing in normal autumn,the field leaf color phenotypes of the low-temperature sensitive material BN044371,the low-temperature sensitive material Bainong4199,and Bainongaikang58 were tracked.Compared with Bainong4199 and Bainongaikang58,the new leaves of BN044371seedlings are normally green in the early autumn season;with the progress of the autumn and winter seasons,the temperature drops to above 0?,about 4?,the new leaves will appear The phen omenon of yellow spots;as the temperature rises in spring,the new leaf tissue will return to green as the temperature increases.In contrast,the low-temperature-sensitive materials Bainong4199 and Bainongaikang58 remained normally green throughout the seedling period.Therefore,the optimal t emperature for the identification of low-temperature susceptibility is above 0?,about 4?.2.Genetic analysis of leaf low temperature sensitivityGenetic analysis was conducted on the F₁,F₂,and F_2:3:3 families of low temperature-sensitive parents BN044371 and its reciprocal crosses constructed with low-temperature-sensitive parents Bainong4199 and Bainongaikang58.The results showed that in the forward and reverse cross exper iments,the leaves of the F1 generation plants all showed the phenomenon of low temperature resi stance;the chi-square values of the separation ratios of the low temperature sensitive individual lea ves and the low temperature sensitive individual plants of the F₂ generation isolated population were all 0.74??²<3.84?,which meets the 3:1 suitability test;the chi-square values of the low-tempe rature resistance coefficient,the separation coefficient,and the low-temperature sensitivity coefficien t of the blades in the F_2:3:3 family of BN044371/Bainong4199 are 1.47 and less than 5.99,which is in line with 1:2:l separation ratio;indicating that the low temperature sensitivity of BN044371 leaves is controlled by a single recessive nuclear gene?wchs1 gene?,the inheritance method is simple,the effect is single,and it is stably inherited in wheat hybrid offspring.3.BSR-Seq analysis resultsFrom the F₂ population of low-temperature sensitive material BN044371 and low-temperat ure sensitive material Bainong4199,30 strains of low-temperature-sensitive wild-type and low-tempe rature-sensitive mutant were selected respectively to construct two mixed sample pools of wild-type and mutant.By sequencing by BSR-Seq method,a total of 384,357 potential SNPs were discovered.The classical Bayesian algorithm was used to locate the mutated gene on the 1DS chromosom e,and on this basis,we independently designed and developed 25 enzyme-amplified polymorphism sequence?CAPS?markers for gene mapping.4.Location of wheat leaf low temperature sensitive gene?wchs1 gene?In this study,180 single plants of F₂ generation and their F_2:3:3 families of low-temperature-sensitive parent BN044371 and the low-temperature-resistant parent Bainong4199 were used as res earch objects.The BSR-Seq method was combined with CAPS molecular markers for analysis.On the wheat 1DS chromosome,two CAPS markers linked to low-temperature sensitive genes were o btained.These two markers were IWGSC and 344678,which were located on both sides of the w chs1 gene,and the genetic distance from the wchs1 gene was 9.8 cM and 12 cM,respectively.