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为了了解黄肉苹果种质成熟时富集类胡萝卜素的分子机制,以黄肉种质‘东北黄海棠’为材料,选择转色期前后(盛花后90 d和115 d)两个发育时期的果实进行RNA-seq测序,分析两个样本差异表达基因,并利用实时荧光定量PCR技术验证获得的类胡萝卜素合成相关差异表达基因的表达水平。结果获得两个样本显著差异表达基因共3 056个,与盛花后90 d果实比较,成熟期果实中有1 270个基因上调表达,1 786个基因下调表达。对这些表达差异基因进行了Pathway富集分析,结果显示差异表达基因涉及类胡萝卜素、苯丙氨酸以及类黄酮等代谢途径,其中类胡萝卜素代谢途径基因有3个上调,4个下调。通过对这7个差异表达基因进一步的qRT-PCR及聚类分析,发现一个新的候选基因MdCCD4b,该基因与菊花以及桃等CCD4聚为一类,与其他作物功能已知的CCD4一样,其表达水平与黄肉种质中类胡萝卜素积累呈负相关,推测黄肉种质果实成熟后类胡萝卜素的富集与MdCCD4b基因显著下调有关。
In order to understand the molecular mechanism of enrichment of carotenoids in germplasm of yellow meat apple, the yellow meat germplasm ’Begonia tuberosus’ was selected as the material to select the two developmental stages The fruits were sequenced by RNA-seq, the differentially expressed genes of two samples were analyzed, and the expression levels of differentially expressed genes related to carotenoid synthesis were verified by real-time fluorescence quantitative PCR. Results A total of 3 056 differentially expressed genes were obtained from the two samples. Compared with 90 days after flowering, 1 270 genes were up-regulated and 1 786 genes were down-regulated in mature fruits. Pathway enrichment analysis of these differentially expressed genes showed that differentially expressed genes involved in the metabolic pathways of carotenoid, phenylalanine and flavonoids, among which three genes were up-regulated and four were down-regulated. Through further qRT-PCR and cluster analysis of the seven differentially expressed genes, a new candidate gene MdCCD4b was found, which was clustered with CCD4 such as chrysanthemum and peach. Like other CCD4 with known functions, The expression of carotenoids in yellow germplasm was negatively correlated with the accumulation of carotenoids in the yellow germplasm. It is speculated that the enrichment of carotenoid in yellow germplasm is related to the significant down-regulation of MdCCD4b gene.