Studies On Idendification Of Genetic Diversity And Cold Tolerance Of Pigeonpea And Its Cold Tolerance Mechanism

Pigeonpea(Cajanus cajan(L.)Millsp)is the sixth largest edible legume crop and also used as animal feed,widely grown in tropical and subtropical regions in the world.The planting area of pigeonpea in China is located in the northern edge of the tropics,where often affected by cold,resulting in a significant decrease in pigeonpea production.The lack of cold-tolerance varieties seriously restricts the development and cultivation of pigeonpea.It is very important to deeply explore excellent cold-tolerance pigeonpea germplasm resources and breed cold-tolerance pigeonpea for improving the production system.In this study,30 pigeonpea core germplasms were obtained from 86 pigeonpea resources by genetic diversity analysis;then the cold tolerance of 30 core accessions was identified,and their different cold tolerance characteristics were gotten;8 pigeonpea materials were used to explore the physiological mechanism of cold tolerance.Finally,the transcriptome and protemic analysis were carried out using the cold tolerance material Miandian(MD)pigeon pea and cold sensitive material Qiongzhong(QZ)pigeonpea,to dissect the molecular mechanism of cold tolerance of pigeon pea.The main results are as follows:1.The genetic diversities of 86 pigeonpea resources were evaluated using 24 phenotypic traits and 42 pairs of SSR markers.A database of phenotypes was established.The diversity index of 10 quality traits ranged from 0.95 to 3.13,and the diversity of seed color was the most abundant.Based on the data of 14 quantitative characters such as flowering period and seed weight,all germplasms were divided into three types: early-maturing with large-grain,middle early maturing with small-grain and late-maturing with medium grain.The 42 pairs of SSR primers showed polymorphism among 86 accessions,the average Shannon diversity index was 0.88,and the average PIC was 0.47,indicating that the tested pigeonpea accessions had rich genetic diversity.The 86 pigeonpea accessions were clustering into 5 groups at the coefficient of 0.557.Based on the results of phenotypic traits and SSR markers,30 pigeonpea core germplasms were obtained from 86 pigeonpea resources.2.The cold tolerance of 30 pigeonpea core germplasms was identified by using relative electrical conductivity and SPAD value under 4℃ stress,and their different cold tolerance characteristics were clarified and 3 cold tolerance pegionpea accessions namely P15,P22 and P35 were also selected.3.8 pigeonpea resources with different cold tolerance were selected and treated under 4°C stress,and REC,contents of SS,MDA,Pro and Chl,activities of SOD,CAT and POD were measured.The characteristics of cold tolerance of 8 attendances were verified by comprehensive evaluation and the sequence from strong to weak as: Miandian > Danzhou > Pigeonpea(170306001)> Pigeonpea(060302216)> ICPL7086 > Nanfan02137 > Guangxi > Qiongzhong.It was found that keeping stable and high content of Chlorophy II and increasing the activities of SOD and POD in enzymatic defense system to enhance cold-resistant ability and alleviate the damage of low temperature stress were the physiological mechanism of pigeonpea cold tolerance.4.Abundant transcription information was obtained by using cold tolerance MD and cold sensitive QZ pigeonpea under 4℃ stress by RNA-seq.A total of 28,796 expressed genes were detected from 12 samples,of which 26,452 were known and 2,344 were new genes predictively.It was found that signal transduction pathway were enriched only in MD with significant difference.There were 150 DEGs in MD different from QZ,which were involved in linoleic acid metabolism,phenylpropanoid metabolism,starch and sugar metabolism,glycerolipid metabolism and etc.All these regulation enhanced their cold tolerance ability.Transcription factors such as MYB,AP2-EREBP,b HLH and WRKY affected their cold tolerance at the same time.5.DIA was performed using MD and QZ as materials.42,769 peptides and 5,794 proteins were identified from 12 samples.DEPs were involved in hydrogen peroxide metabolism,reactive oxygen species and signal transduction,oxidoreductase etc.There were 78 DEPs in MD materials different from QZ response to low temperature stress,which mainly were involved in peroxidases,oxidoreductase,heat shock proteins,amino acid transportases and etc.Signal transduction,over-expressed peroxidases played an important role in cold tolerance materials.In conclusion,the genetic diversities of 86 accessions was abundant.The cold tolerance of 30 core accesions difference and 3 cold tolerance germplasm were gotten.The physiological mechanism of cold tolerance of pigeionpea is to maintain Chl stable high content and enhance SOD,POD activities.The molecular mechanism of cold tolerance of pigeonpea may be the regulation of signal transduction,linoleic acid metabolism,phenylpropane biosynthesis,starch and sugar metabolism and glycerol metabolism pathway,and improve the expression of peroxidases to adapt to the cold stress.This study laid an important foundation for the cold tolerance germplasm breeding of pigeonpea.