| Chinese cabbage is a very important cruciferous vegetable crop with the largest acreage in China.It has a long history of cultivation,rich variety and distinct morphological characteristics.The leaf is both a photosynthetic organ and a product organ,providing a place for photosynthesis.Therefore,the related research on the metabolic pathway of leaf color mutation gene in Chinese cabbage is particularly important.Leaf color mutants are valuable resources for studying the regulation mechanism of photosynthetic pigment metabolism.In this study,a chlorophyll-deficient golden leaf mutant lcm1 of Chinese cabbage was identified from its wild-type DH line 'FT' by EMS mutagenesis,its photosynthetic physiological characteristics were confirmed,and candidate mutation sites were identified.The main findings are as follows:1.After planting and identification in spring and autumn,it was found that the golden leaf mutant lcm1 showed whole plant yellowing during the whole growth period,and its mutant traits were not affected by external environmental factors,and the stalks,flower buds and pods of the mutant appeared different degrees of yellowing.It can be seen from the analysis of the growth amount of the seedling mutant and the wild type that the dry weight of the golden leaf mutant lcm1 was significantly lower than that of the wild type 'FT' throughout the observation period.2.By measuring the photosynthetic pigment content of the mutants in the four-leaf stage and the wild type,it was found that the total chlorophyll,chlorophyll a and chlorophyll b contents of the leaves of the chlorosis mutant lcm1 were significantly lower than those of the wild type 'FT' and the carotenoid content of the lcm1 was only slightly lower than the wild type.3.In order to study whether the yellowing of Chinese cabbage is related to the damage of chloroplast structure,the ultrastructure of the golden leaf mutant lcm1 and the wild type ‘FT' of Chinese cabbage was observed by transmission electron microscopy.The results showed that the chloroplast ultrastructure of the wild type 'FT' and the mutant lcm1 was significantly different.The chloroplast structure of the wild type is regular and full,the thylakoids are tightly packed,and the granule structure is obvious.The accumulation of starch granules also can be seen.The thylakoid of the mutant lcm1 is only maintained as a single original layer,no obvious granule structure exists,and no obvious accumulation of starch granules is observed.4.Genetic analysis indicated that the mutant yellowing trait was controlled by a single recessive nuclear gene lcm1(BrChlH).The MutMap method and Kompetitive Allele Specific PCR genotyping were used to predict the candidate gene for the mutant lcm1.A SNP was found that co-segregated with the mutant phenotype and located on exon 3 of Bra006208 at 2523361 on chromosome A03.The base at the site is G in the wild type,and A in the mutant,resulting in the amino acid being changed from A(alanine)to V(valine), and the gene encoding the Magnesium protoporium IX chelate enzyme H subunit.This SNP2523351 occurred in the highly conserved CobN-Mg_chel domain of the LCM1(BrCHLH)protein.Therefore,Bra006208 was used as a candidate gene for lcm1.5.Transcriptome analysis showed that there were 606 differentially expressed genes in wild type and mutant leaves,which were mainly enriched in metabolic pathways such as Biosynthesis of secondary metabolites and Carbon fixation in photosynthetic organisms.By sequencing the clones of the wild type and mutant regions,it was found that the SNP mutation site was actually present.This study suggests that the amino acid at the site of the mutant SNP plays an important role in maintaining the function of the magnesium-chelatase subunit chlH and lcm1 gene plays an important role in the function of CHLH and provides a solid foundation for further studies on the development of plant leaf color... |
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