Change Patterns Of Hormones And Quality Characters During Taproot Growth And Development In Carrot

Carrot (Daucus carota L.),a biennial plant of Apiaceae family,is an important root vegetable worldwide. Fleshy taproot is the main edible part and its growth and development directly determines the yield and quality of carrot. Fleshy root undergoes a complex process that involves anatomical changes, biomass accumulation, and gene regulation, which is under tight control of plant hormones. However, hormone accumulation and its molecular mechanisms during carrot growth and development remain unclear. Meanwhile, reports regarding ascorbate and lignin accumulation from the point of view of molecular biology have not been identified. In the present study, high-throughput sequencing was introduced to identify differential expression genes during carrot taproot development, and hormone-related genes were selected and further analyzed to elucidate the potential roles of hormones in carrot growth. Three different software tools (NormFinder,geNorm,and BestKeeper) evaluating expression stability of candidate genes were applied to select the optimum reference gene(s) for developmental processes in carrot. We also analyzed the accumulation of hormones, ascorbate, and lignin at different developmental stages based on morphological, physiological, anatomical, cellular, and molecular methods. In addition, the effects of exogenous gibberellin on taproot growth and quality characteristics were studied.The major findings are as follows:1. We collected carrot root samples from four developmental stages, and performed transcriptome sequenceing to understand the molecular functions of plant hormones in carrot. A total of 160,227 transcripts were generated from our transcriptome, which were assembled into 32,716 unigenes with an average length of 1,453 bp. A total of 4,818 unigenes were found to be differentially expressed between the four developmental stages.In total, 87 hormone-related differentially expressed genes were identified, and the roles of the hormones are extensively discussed. The results suggest that plant hormones may regulate carrot root growth in a phase-dependent manner.2. Three different software tools including NormFinder, geNorm, and BestKeeper were applied to detect expression stability of nine candidate genes (ACTIN, EF-1?, eIF-4?,GAPDH, PP2A, SAND, TIP41, TUB and UBQ). The results showed that the best reference genes differed across the tissues and developmental stages. ACTIN was recommended as the most stable gene in carrot roots and leaves at five developmental stages. GAPDH appeared to be the least adequate gene indicating that it should be avoided as a reference gene in carrot. Thus, we suggest ACTIN or different combinations of ACTIN, EF-1?, eIF-4?,TUB, and UBQ for normalization in carrot developmental processes.3. Carrot plants at five stages were investigated using morphological and anatomical structural techniques. Gibberellin, cytokinin, and MeJA levels in carrot tissues were also investigated for all five stages. Genes involved in hormone biosynthesis and signaling were identified from CarrotDB, and their expression levels were determined. All of the genes were evidently responsive to carrot growth and development, and some of them showed tissue-specific expression. However, transcript levels of most genes did not correlate well with hormone levels, indicating that hormone accumulation and signal transduction was regulated through a complex network in carrot. The results suggested that gibberellins,cytokinins, and jasmonates may play vital roles in carrot growth.4. To gain insights into the regulation of AsA accumulation and to identify the key genes involved in the AsA metabolism, carrot variety 'Kurodagosun' was used in this study.We cloned and analyzed the expression of 21 related genes during carrot root development.The results indicate that AsA accumulation in the carrot root is regulated by intricate pathways, of which the L-galactose pathway may be the major pathway for AsA biosynthesis. Transcript levels of the genes encoding L-galactose-1-phosphate phosphatase and L-galactono-1,4-lactone dehydrogenase were strongly correlated with AsA levels during root development.5. We collected taproots of wild and cultivated carrots at five developmental stages and analyzed the lignin content and characterized the lignin distribution by using histochemical staining and autofluorescence microscopy. Genes involved in lignin biosynthesis were identified, and their expression profiles were determined. Results showed that lignin was mostly deposited in xylem vessels of carrot root. In addition, lignin content continuously decreased during root development, which was achieved possibly by reducing the expression of the genes involved in lignin biosynthesis.6. Carrot roots were treated with GA3 and the effect of applied GA3 on plant growth,particularly root growth, xylem development, and lignin accumulation was extensively studied. The results indicated that GA treatment dramatically stimulated the shoot growth but reduced the root growth of carrot. Significant cell wall thickening was also detected in the xylem parenchyma. Autofluorescence analysis with UV excitation indicated that these cells became lignified due to the long-term GA3 treatment. Transcript levels of genes related to gibberellins, auxin, cytokinins, abscisic acid and brassinolides were altered in response to increased gibberellins. Moreover, lignin content was found to be increased in carrot root, and transcripts of lignin biosynthetic genes were altered in response to applied GA3. The results indicated that GA may play important roles in xylem growth and lignification in carrot roots.