Cloning And Expression Analysis Of α-Galactosidases In Cucumber (Cucumis Sativus L.)

Cucumber(Cucumis sativus L.) primarily transports stachyose in the phloem, while in the fruit, the predominant soluble sugars are sucrose, glucose and fructose, little raffinose family oligosaccharides can be detected in this tissue. Nevertheless, the details how stachyose unloaded from the phloem and then converted to sucrose still remains poorly understood by far. Investigating the expression pattern ofα-galactosidase (EC 3.2.1.22), a key enzyme catalyzing stachyose catabolism, at tissue, cellular, and subcellular level is beneficial to answer these questions. Cloning the cDNA sequences ofα-galactosidases in cucumber is the first step to suit this purpose.In this study, 4 full length cDNA sequences ofα-galactosidases were obtained by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and Rapid Amplification of cDNA Ends (RACE) in cucumber, according to homological sequences ofα-galactosidase genes in other plants. They are named: cucumber acidα-galactosidasesⅠ(DQ320569), cucumber acidα-galactosidasesⅡ(DQ361277), cucumber alkalineα-galactosidasesⅠ(DQ157703) and cucumber alkalineα-galactosidasesⅡ(DQ395330). To our knowledge, this is the first report of cloning full length cDNAs of 2 acid and 2 alkalineα-galactosidases in the same plant. Thus, we can design probes, primers or RNA interference based on these sequences to do further research work in this area.The expression patterns of mRNAs of theseα-galactosidases were inspecteded by RT-PCR. The results showed that except dry seed, 4 genes are expressing in different tissues, including peduncle, petal, stamen, stem, hair roots and fruit at different developmental stages in cucumber. These mRNAs were also detected in seed during germination. The data indicated that all 4α-galactosidases we studied in this paper play important physiological roles in cucumber.