Comprehensive Evaluation Of Salt And Alkali Tolerance And Transcriptome Analysis Lotus

In our country,the area of saline-alkali land with different degrees is increasing year by year.It is of great theoretical significance and application value to explore the salt-alkaline tolerance of plants to expand the planting area of saline-alkali land and improve the ecological environment.Lotus,as one of China’s traditional famous flowers and important aquatic plants,has important ornamental and economic value.In this paper,the method of saline-alkali stress was used to set the concentration of 100 mM mixed saline-alkali(NaCl:NaHCO3=2:1)solution and control for treatment.After the test for seven days,according to the comprehensive analysis of fourteen morphological and physiological indicators,the salt-tolerant of twenty lotus resources were tested.The evaluation and classification of salt-tolerant varieties and salt-sensitive varieties were carried out before and after the salt-alkali stress treatment by transcriptome sequencing and analysis.The Preliminary reveals the mechanism of lotus salt-tolerance from the molecular level,and provides valuable scientific and theoretical basis for the resistance breeding of lotus and other garden plants.The main research results are as follows:(1)The effect of salt-alkali stress on the growth and physiology of lotus.The potted method was used to treat twenty kinds of lotus with 0 mM,100 mM saline-alkali,and the morphological indexes of lotus after seven days of saline-alkali stress treatment:plant height,leaf area;physiological indexes:antioxidant enzyme activity,osmotic adjustment substance content and malondialdehyde content.The results showed that under saline-alkali stress,in addition to the ratio of chlorophyll a,b,malondialdehyde,proline,soluble protein and soluble sugar,plant height,leaf area,chlorophyll a content,chlorophyll b content,total chlorophyll content,PS Ⅱmaximum photochemistry The index values of efficiency and potential activity of PS Ⅱ decreased under saline-alkali stress.(2)Comprehensive evaluation of the salt-alkali tolerance of lotus.Through principal component analysis and cluster analysis,twenty kinds of lotus are divided into four grades according to their salt and alkali resistance:salt and alkali resistance materials(four parts),medium salt and alkali resistance materials(five parts),weak salt and alkali resistance materials(eight parts),sensitive salt-alkali materials(thirteen parts).Establish a mathematical model for evaluating the salt and alkali tolerance of lotus.D=0.453+0.148PH+0.122LA-0.122MDA-0.033Pro+0.093SS+0.037SP,it shows that the six indexes of plant height(PH),leaf area(LA),malondialdehyde(MDA),proline(Pro),soluble sugar(SS)and soluble protein(SP)have significant effects on the salt and alkali tolerance of lotus;the estimation accuracy of the equation is all above 95.45%,indicating that the equation can be used to evaluate the salt-alkali tolerance of lotus.(3)Lotus transcriptome analysis.Using high-throughput sequencing technology,using saline-tolerant varieties‘HuangShuai’and sensitive saline-alkali varieties’TaiWanMoPanLian’ as test materials,a total of twelve samples of leaf tissues of two materials treated with 0 mM and 100 mM saline-alkali stress for seven days were sequenced.After data assembly,a total of 146786 Unigenes were obtained,with a total length of 93,817,868 nt and an average length of 639.15 nt.The comparison in nine public databases showed that the number of genes successfully annotated by 146786 Unigenes in at least one database was 84669,and a total of 101 transcription factors were obtained,which were mainly divided into thirteen transcription factor families.(4)Differential gene and enrichment analysis.The results showed that a total of 146,786 Unigenes were obtained,and there were a total of 4916 DEGs among the four samples,including 3101 DEGs up-regulated expression and 1815 DEGs down-regulated expression.There were 117 DEGs in salt-alkali-tolerant varieties under saline-alkali stress,and 652 DEGs in salt-sensitive varieties under saline-alkali treatment.In the KEGG analysis,the salt-tolerant materials DEGs were mainly enriched in phenylpropionic acid biosynthetic metabolic pathways,followed by phenylalanine metabolism,plant MAPK signaling pathways and plant pathogen interaction pathways.DEGs of sensitive saline-alkali materials after saline-alkali stress are mainly concentrated in metabolic pathways such as carbon metabolism,phenylpropionic acid biosynthesis and carbon fixation in the photosynthetic system.This indicated that the phenylpropionic acid biosynthetic metabolism pathway and plant MAPK signal pathway under saline-alkali stress may be important reasons for the salt-alkali-tolerant varieties to adapt to the salt-alkali stress environment.Real-time fluorescence quantification(qRT-PCR)was used to verify the expression of twelve randomly selected DEGs.The result is that the relative expression level is basically consistent with the transcriptome sequencing result,indicating that the sequencing data is true and reliable.