Leaf Color QTLs Mapping In Peanut

The color of plant leaves is the comprehensive embodiment of various pigments in leaves,which is the result of the combined action of internal and external factors.In higher plants,the pigments that affect the color of leaves mainly include chlorophyll,carotene and anthocyanin,and the proportion of these pigments and selective absorption of light lead to the difference of leaf color.Plant leaf color variation is a common mutant character,which directly affects photosynthesis and has an important influence on the growth and development of plants and crop yield.With the rapid development of genomics,bioinformatics and new molecular biology technologies,more and more leaf color related genes have been characterized,and leaf color mutants have been widely used.,As an important cash crop and oil crop,the leaf color of peanut(Arachis hypogaea L.)plays an important role in the formation of peanut yield and quality.The mechanism of leaf color variation in major crops such as rice and wheat has been studied deeply,while the research on peanut is still limited.It is not only restricted the use of peanut germplasm resources,but also seriously affects the molecular breeding process of peanuts.Therefore,mapping QTLs/genes controlling leaf color traits in peanut and revealing the molecular mechanism of leaf color variation are of great significance to the breeding of peanut varieties with high yield and good quality.In recent years,whole-genome sequencing of the diploid ancestor and cultivars of peanut has been completed,and our research group has developed whole-genome SSR markers,which promoted the molecular analysis of important agronomic traits in peanut.In this study,BSA(Bulked Segregant Analysis)sequencing,physiological index detection,expression profile analysis and other methods were used to identify the QTLs and genes related to peanut leaf color,which laid a foundation for the mining and molecular Analysis of peanut leaf color trait genes.The main results are as follows:(1)In order to explore which abnormal pigment content caused the change of peanut leaf color,we measured the contents of chlorophyll,carotenoid and anthocyanin in the leaves of 3 yellow leaves and 3 green leaves phenotypic plants from the parents and offspring groups.The results showed that the contents of chlorophyll,carotenoid and anthocyanin in green leaf plants and parent Tifrunner were significantly higher than those in yellow leaf plants and parent ID.We speculate that the decrease of chlorophyll content is the main reason for the yellow color of leaves,while anthocyanin may be the reason for the green leaves to be more green.(2)Based on the survey of leaf color separation data in F2 segregation population,25 green and 25 yellow single plant of F2 generation with obvious leaf color differences in the growth period were selected,and the leaf DNA was extracted.Two extreme pools of yellow and green leaf phenotypes were constructed by mixing the same amount of DNA.Perform BSA sequencing.The results showed that the differential genes were mainly distributed on chromosomes 02,08,10,and 12,followed by shorter intervals on 01,04,05,07,and 09 chromosomes.(3)Validation of the BSA results with SSR and InDel molecular markers resulted in 10 pairs of SSR primers designed on chromosome 2 spanning 84.7 Mb-97 Mb.SDS-PAGE gel of their amplified products resulted in one SSR marker that were polymorphic and linked to leaf color.The marker was located at 95,935,239 bp on chromosome 02 and named A02-95.Six pairs of InDel primers were designed in the mapping region of chromosome 08.There are two SSR markers that were polymorphic and linked to leaf color,named A08-2 and A08-4.In order to analyze the linkage relationship between these primer and the leaf color characters of peanut,8pairs of primers were designed and synthesized in and near the area where the marker is located.Among them,three markers are polymorphic and have a linkage effect with leaf color.These 3 marks are respectively named A02-95-2,A02-96-1,A02-96-4.14 pairs of primers were designed and synthesized between two markers on chromosome08.There are three markers were polymorphic and linked with leaf color.The three markers were named A08-16,A08-22 and A08-24.(4)The 4 markers developed on chromosome 02 were tested in 50 yellow and 52 green F2 individuals of the population.The genetic map and QTL were constructed by using the software of QTL icimapping 4.2.The results showed that a QTL related to peanut leaf color was detected at this location,which was located between the markers a02-96-1 and a02-96-4.The physical locations of the two markers were 96.17 Mb and 96.98 Mb,respectively,with a distance of 0.81 Mb.This leaf color regulatory gene locus can explain 11.86 % of the phenotypic variation rate.(5)In order to further analyze the molecular mechanism of peanut leaf color gene regulation,we analyzed the expression profile of yellow leaves and green leaves,combined with BSA analysis results,to screen genes that may be involved in the regulation of leaf color.The results showed that the expression of heme related genes was up-regulated,the expression of magnesium ion transport related genes,photosynthesis protein related genes and lipoxygenase genes down-regulated,which would cause the decrease of chlorophyll content and the relative content of carotenoids,leading to the production of leaf yellow phenotype.