A Preliminary Study On The Physiological Functions Of Sixα-galactosidases In Cucumber (Cucumis Sativus L.)

Six α-galactosidase genes exist in the cucumber(Cucumis sativus L.) genome. The results of preliminary studies and sequence analysis showed that there are three alkaline forms and three acidic forms. The physiological functions of these alpha-galactosidases during cucumber growth and development still remains poorly understood so far. Real-time quantitative PCR was used to study these a-galactosidase gene expression patterns. On the basis of studying the relative expression of the six genes in different organs, possible physiological functions of these enzymes during the fruit development, seed germination, leaf development and senescence were suggested. The results showed that:1、Product sequencing and dissolution test showed that the primer we used can guarantee the specificity of real-time PCR. From the standard curve we can see the efficiency of18SRNA and6a-galactosidases are all close to1. The result of Northern blot showed that the real-time PCR system we established is proper for the study of expression level of6a-galactosidase genes.2、All kinds of a-galactosidase genes are expressed in cucumber roots, stems, leaves, flowers, fruits. The expression level of the alkaline a-galactosidase genes are higher than that of acidic a-galactosidase genes.The relative expression level of the acidic form Ⅰ gene is higher in cucumber leaves. The relative expression level of acidic form Ⅱ gene is very low in cucumber’s root, male flowers and female flowers. In male flowers and female flowers the expression ratio of the acidic form Ⅲ gene is also very low. The alkaline a-galactosidase Ⅱ has a higher expression in all the other organs of cucumber except male flowers. In the male flowers, the expression of the alkaline a-galactosidase enzyme Ⅰ is highest. These results suggest that alkaline a-galactosidase Ⅱ may play an extensive and important role in the process of cucumber growth and alkaline form Ⅰ may have important functions in male flowers.3、Compared to the expression level of pedicel, three kinds of acidic a-galactosidases showed little changed and alkaline a-galactosidase Ⅰ and Ⅲ displayed high level in the apex flesh and placenta vascular. They may play a major role in fruit growth and seed development. Compared with pedicel, the expression of alkaline form II decline with the change of the RFOs content in the fruit, suggesting that it may be involved the unloading of the stachyose. 4、The expression level of alkaline form Ⅱ and acid form Ⅱ in the young leaf remains high and then declined when the leaf reach the mature stage, implying that alkaline Ⅱ may be involved in the stachyose unloading to the sink leaf tissue, while the acid Ⅱ may play a role in the young leaf growth; the alkaline Ⅲ and acidic Ⅲ expression in senescent leaves was significantly higher than that in the mature leaves, suggesting they may be involved in the decomposition of polysaccharides in senescent leaves of cucumber.5、According to the paraffin section and the changes of the soluble sugar contents during seed germination, alkaline form Ⅱ may play a role in the degradation of RFOs at the early germination stage. While other forms of a-galactosidase may be involved in the polysaccharide decomposition, cell division and cell enlargement at the later germination stage.