番茄果胶裂解酶基因SlPL参与调控裂果机制研究

Study on the Regulatory Mechanism of SlPL Gene Affecting Tomato Fruit Cracking

以裂果率差异极显著的番茄为材料,分析了SlPL(Solyc03g111690)基因的表达差异,进一步利用基因遗传转化对其进行功能验证。结果表明,SlPL在易裂番茄‘NT189’中的表达量显著高于耐裂番茄‘NT91’;在番茄果实中的表达量显著高于根、茎、叶、花等器官,且在果实转色期和红熟期表达量较高;通过灌水处理和ABA处理诱导果实开裂,发现在相同处理时期,易裂果材料果实中SlPL表达量总体显著高于耐裂果材料。通过遗传转化获得SlPL过表达(OEPL)和敲除(pl)的株系,与野生型相比,OEPL更易裂果,且果实硬度显著降低,pl果实硬度升高。OEPL果实中原果胶含量显著低于野生型,水溶性果胶含量显著高于野生型,pl果实中原果胶和总果胶含量显著高于野生型;OEPL果实中果胶裂解酶活性显著高于野生型,pl果实中果胶裂解酶活性显著低于野生型。基因表达分析发现,OEPL果实中细胞壁代谢相关基因SlPG2、SlPME2.1、SlCel2、SlGH9C5和乙烯合成途径相关基因SlACS4、SlACO1的相对表达量显著高于野生型,pl果实中则相反。pl果实中乙烯响应因子SlERF2的相对表达量显著高于野生型。果皮显微结构观察发现,与野生型相比,OEPL表皮层细胞和薄壁细胞排列稀疏,而pl果皮细胞排列更为紧密。

Tomato with highly significant differences in fruit cracking rate were used as the materials to analyze the expression differences of the SlPL(Solyc03g111690)gene,which was further functionally validated using genetic transformation.The results showed that the expression of SlPL was significantly higher in crack-susceptible tomato‘NT189’than in crack-resistant tomato‘NT91’;in tomato fruit was significantly higher than that in other organs such as roots,stems,leaves and flowers,and the expression was higher in the break ripen and red ripen stages of the fruit.Fruit cracking was further induced by irrigation and ABA treatments,and it was found that the expression of SlPL was overall significantly higher in the fruit of crack-susceptible materials than that of crack-resistant ones in the same treatment period.SlPL overexpression(OEPL)and knockout(pl)lines were then obtained by genetic transformation.OEPL was more susceptible to fruit cracking and had significantly lower fruit firmness and pl had higher fruit firmness than the wild type.OEPL fruit had significantly lower pro-pectin content than the wild type,and significantly higher water-soluble pectin content than the wild type,while pl fruit had significantly higher pro-pectin and total pectin content than the wild type.Pectin cleavage enzyme activity was significantly higher in OEPL fruits than in wild type,and pectin cleavage enzyme activity was significantly lower in pl fruits than in wild type.Gene expression analysis revealed that the relative expression of cell wall metabolism-related genes SlPG2,SlPME2.1,SlCel2,SlGH9C5,and ethylene synthesis pathway-related genes SlACS4 and SlACO1 was significantly higher in OEPL fruits than in wild type,and the opposite was true for pl fruits.pl fruits showed a significantly higher relative expression of the ethylene-responsive factor SlERF2 than in wild type.Observations of the pericarp microstructure revealed that the cells of the epidermal layer and thin-walled cells were more sparsely arranged in OEPL compared with wild type,and those cells were tightly arranged in pl.