Comparative Transcriptome Analysis Of 'Dongzao' Jujube And 'Jinsixiaozao' Jujube In Response To Freezing Stress

Jujube(Ziziphus jujuba Mill.)is a dried fruit tree species native to China.It is loved by consumers.In recent years,global climate warming has led to frequent occurrence of extreme weather events,resulting in frequent occurrence of jujube freezing injury,which has brought great losses to jujube production.Breeding strong fold resistant varieties is an important way to solve this problem.Therefore,breeding for frost resistance has become an important goal of jujube breeding.This study compared the results of frost resistance measurement by three methods,i.e.direct observation method,TTC staining method,and relative electrolyte leakage.The relative electrolyte leakage was selected to determine the frost resistance of jujube.Two varieties('Dongzao' jujube and 'Jinsixiaozao' jujube)with significant difference in frost resistance were screened out,and the important genes and metabolic pathways related to frost resistance of jujube were revealed through transcriptomic analysis,in order to provide the basis for the study of frost resistance molecular mechanism and frost resistance breeding of jujube.The main results are as follows:(1)Comparing the three methods for determining the frost resistance of jujube showed that TTC staining method and the direct observation method were simple and low cost,however,the TTC staining method can not distinguish the dyeing levels of samples after different temperatures.The calculated LT50 is very different from the actual LT50 of the varieties,and the measurement result is inaccurate,the direct observation method has the disadvantage of subjective test error,and the relative electrolyte leakage compared with the TTC staining method and the direct observation method,although there are many test procedures and the cost is slightly higher,but the measurement results are accurate,the error is small.Relative electrolyte leakage was used to determine the frost resistance of 30 jujube varieties,the frost resistance of 'Jinsixiaozao'jujube was the strongest,the LT50 was-40.314?,the frost resistance of'Dongzao'jujube was weak,the LT50 was-33.471?,they were selected as materials for transcriptome sequencing.(2)Transcriptome sequencing the xylem of one-year-old shoots of 'Dongzao' jujube and'Jinsixiaozao' jujube at 4?,-10?,-20?,-30? and-40?,produced a total of 144.85Gb data.1831,2030,1993,1845 and 2137 significant differentially expressed genes were identified under each temperature treatment.Among them,854 genes were common under different freezing stress treatments.Differential expression of these genes may be an important cause for the difference in frost resistance of'Dongzao' jujube and 'Jinsixiaozao' jujube.Ten differentially expressed genes related to frost resistance were selected for RT-qPCR,the results showed that the expression levels of 'Dongzao' jujube and'Jinsixiaozao' jujube were basically the same as that of the transcriptional analysis.GO analysis of these significantly differentially expressed genes between 'Dongzao' jujube and 'Jinsixiaozao jujube revealed that these are mainly enriched in the metabolic process,cellular process,cell,cell part,catalytic activity,binding related to biological functions.In the KEGG metabolic pathway analysis,the difference genes between 'Dongzao' jujube and 'Jinsixiaozao' jujube are mainly enriched in Biosynthesis of secondary metabolites,Fructose and mannose metabolism and Biosynthesis of amino acids.(3)The four types of alternative splicing were analyzed:Exon skipping;Intron retention;Alternative 5'splice site;Alternative 3'splice site,with the largest number of alternative splicing is Intron retention,and the number of alternative splicing of 'Jinsixiaozao'jujube is significantly more than the number of alternative splicing of'Dongzao'jujube.