| Camellia sinensis is an important economic crop in China and one of the top three drinks in the world.In the breeding work,it was found that the polyploid tea plant has obvious advantages that the diploid can not be compared.The morphology and physiology of the plant have a series of obvious changes with the increase of the ploidy of the chromosome composition in the nucleus,especially in the growth.Many physiological and developmental processes,such as rate,genetic gain,and increased metabolite yield,are affected by increased ploidy.In this study,the second-generation transcriptome sequencing technique was used to study the growth and development of tea diploid and triploid leaves from the diploid ‘Qian Mei 419' and the triploid ‘Qian Fu 4'.The morphological and physiological differences were analyzed.The molecular mechanism of the growth and development of diploid and triploid leaves of tea trees can reveal the changes in molecular level during the growth and development of leaves and the regulation of key genes,and enrich the theoretical basis of molecular biology of plant organs.On the other hand,the study of the molecular mechanism of its leaf growth and development can provide a theoretical basis for its molecular breeding,breeding excellent new varieties and solving practical problems in production.The main results are as follows:Under field growth conditions,the leaves of triploid tea trees had obvious growth advantages compared with diploid tea leaves,which increased by 23.37%,41.12% and 59.81%,respectively,compared with diploid leaf length,leaf width and leaf area.Significant difference.The stomata size(length and width)of the triploid leaves was significantly larger than that of the diploid,which was increased by 57.20% and 84.44%,respectively,compared with the diploid length and width.However,the number of stomata in triploid leaves is significantly lower than that in diploid,and the number of stomata in diploid tea leaves is twice that of triploid stomata.In short,the diploid leaves have larger stomata,but the density is smaller than the diploid.The xylem of the triploid vein is more developed than the diploid,and the area of the triploid xylem is three times that of the diploid,which is 0.476 mm2.The number of xylem cell layers increased,and the number of triploid xylem cells increased by 30.67% compared to diploid.The shape and size of the cells of diploid and triploid mesophyll were significantly different.The epidermal cell thickness of triploid mesophyll was 22.28% larger than that of diploid,which was significantly different.The cells of the leaf bark of diploid tea leaves are arranged closely,and the shape and size are relatively uniform.The cells of the triploid leaf mesophyll puncture are loosely arranged,the cell gap is large,and the cell shape and size are different.The average length of the diploid palisade tissue cells is larger than that of the triploid,and the length is 65 ?m,which is 15.65% longer than the triploid.The width of the triploid palisade tissue cells was significantly larger than that of the diploid,which was 70% higher than that of the diploid.Diploid sponge tissue cells are small and dense,and the number is more than that of triploids,which is about twice the number of triploid cells.Analysis of secondary metabolites in diploid and triploid tea leaves by high performance liquid chromatography.The results showed that the caffeine content of triploid tea was significantly higher than that of diploid,which was 56.14% higher than that of diploid,and the content reached 1.78% of the dry weight of tea leaves.The content of theanine was significantly decreased.Compared with diploid,the content of theanine in the leaves of triploid tea decreased by 31.34%,and the content of triploid theanine was 1.005g/Kg.Triploid tea leaves catechins(Catechin,C),epicatechin(EC),epigallocatechin(EGC),epicatechin gallate(ECG)and The content of epigallocatechin gallate(EGCG)was significantly lower than that of diploid,and the contents were 0.035%,0.135%,0.22%,0.64%,and 0.645%,respectively,relative to diploid reduction.53.33%,44.90%,33.33%,30.43%,29.12%.Among them,Catechin(C)and Epicatechin(EC)were the most obvious.Epigallocatechin gallate(EGCG)was the highest in diploid and triploid tea leaf catechin monomers,and reached 0.91% and 0.645% in dry leaves.The transcriptome sequencing analysis was used to analyze the difference in leaf gene expression between diploid and triploid of tea tree.A total of 7534 genes were up-regulated and 16459 genes were down-regulated.The transcriptome differential genes were analyzed by combining the morphological and physiological differences between diploid and triploid tea leaves.The results showed that the gene expression of photosynthesis photoreaction stage of diploid tea tree showed a down-regulation trend,while the expression level of related genes in the dark reaction stage was up-regulated.The overall transcriptional and cell division key enzyme genes of triploid are up-regulated.The increase of chromosome multiples in triploid tea tree changes the key enzymes in the secondary metabolite synthesis pathway,which affects the biosynthesis of secondary metabolites in tea plants.Caffeine synthase(TCS 1),a key enzyme regulating gene for secondary metabolites in tea plants,was up-regulated.Theanine synthase(TS)gene is down-regulated.The anthocyanidin reductase(ANR)gene and the serine carboxypeptidase-like acyltransferases(SCPL)gene are key enzymes for the synthesis of catechin monomers EC,EGC,ECG and EGCG.The ANR and SCPL genes were down-regulated.From the level of gene expression regulation,the changes in the content of secondary metabolites and the expression levels of key enzyme genes that regulate the synthesis of secondary metabolites are consistent.The stomata of triploid leaves are larger than diploids,but the density of stomata is significantly reduced.The transcriptome analysis of key genes regulating stomatal development found that B.subtilis serine protease SDD1 negatively regulated the stomatal development of triploid tea leaves,which reduced the stomatal density of triploid tea leaves.The functional analysis of the serine/threonine protein kinase FRAY2 gene transgenic tobacco showed that the growth and development of transgenic tobacco leaves were significantly higher than that of wild type,but there was no significant change in the growth of tobacco roots.Statistical analysis of tobacco leaf size was started on day 0 after transplanting.From the 10 th day,although the leaves are larger than the wild type,the growth potential is significantly reduced.Therefore,it is hypothesized that the FRAY2 gene plays an important regulatory role only in the early stage of leaf growth and development. |
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