关键词: 罗汉果, 葡萄糖基转移酶, 基因克隆, 原核表达, 时空表达

Abstract: A glucosyltransferase gene，named as SgUGT4，was cloned from Siraitia grosvenorii fruits at 70 days after pollinating（DAP）by RT-PCR and RACE methods. The full-length cDNA of SgUGT4 is 1 726 bp and has an open reading frame（ORF）of 1 344 bp which encodes a predict protein of 447 amino acids. The recombinant plasmid pET32a-SgUGT4 was constructed in a prokaryotic expression system and then was transformed into E.coli strain Rossetta-gami（DE3），A 65 kD fusion protein was expressed after being induced by IPTG. Phylogenetic tree analysis showed that SgUGT4 was classified as the same subfamily as the glucosyltransferase of AtUGT73C3，AtUGT73C5，AtUGT73C6 from Arabidopsis thaliana and SrUGT73E1 from Stevia rebaudiana，which could catalyze mogrosides biosynthesis.qRT-PCR analysis demonstrated that the expression level of SgUGT4 in pulps where mogroside Ⅴ most abundantly accumulated was more higher than that in stems，leaves，flowers and peels. SgUGT4 did not nearly expressed in roots and seeds. From 40 DAP to 50 DAP，the content of mogrosideⅤgradually occurred and accumulated，the expression level of SgUGT4 rose gradually. The expression level of SgUGT4 up-regulated sharply after 50 DAP. The more higher the mogroside Ⅴ content in varieties was，the more higher its expression level of SgUGT4 in varieties was. Therefore，the SgUGT4 probably involved in mogroside Ⅴ biosynthesis.

Key words: Siraitia grosvenorii, glucosyltransferase, gene cloning, prokaryotic expression, temporal and spatial expression