The Full Length cDNA Cloning of Persimmon ADH Gene and Its Expression Analysis in Astringent and Non-Astringent Fruits

摘要

Alcohol dehydrogenase (ADH, EC 1.1.1.1) plays a role in plant development, stress responses and metabolite synthesis. In persimmon fruit, ADH is closely related to fruit development and de-astringency of harvested fruit. The full length of ADH cDNA was cloned from 'Mopanshi' persimmon (Diospyros kaki) fruit, using RT-PCR and RACE techniques with degenerate primer which were designed according to the conservative sequences (DVYFWEA and DHVCEFA) of dicotyledon ADH. Persimmon (Diospyros kaki 'Mopanshi')fruit was used as experiment material to clone a full-length clone of ADH cDNA sequence of persimmon. The sequence and structure analysis of the ADH, and its spatial and temporal expression in fruits of different persimmon cultivars ('Jirou' and 'Mopanshi', PCNA- and PCA-cultivar, respectively) were also studied. The full sequence of ADH cDNA was 1459 bp in length, namely DkADHX, and deduced molecular weight of 41 kDa, whose open reading frame (ORF) was 1140 bp in length, encoding 379 amino acid residues. Results of real-time PCR analysis showed that the ADH expression differed in relative expression amount between, 'Mopanshi' (a PCA type) and 'Jirou' (a PCNA type fruit). The relative expression level in peel was significantly higher than in pulp, in-4.5 times. The relative expression level in 'Jirou' fruit was high during July-September, then, dropped in October, however, in 'Mopanshi' fruit, the expression dropped gradually from July to October. Results indicated that the ADH gene expression in persimmon fruit was related to type of fruit, fruit development stages, and fruit tissues.