关键词: 瓯柑, 中性转化酶, 蔗糖合成酶, 基因克隆, 基因表达

Abstract: The purpose of this work was to separate NIN and Sus gene from Ougan fruit and analyze expression patterns of both two genes in different tissues at different development stages of fruits， which will provide some basic information for sugar metabolism of Ougan fruit and enrich NIN and Sus study on citrus fruit. Two full length sequences encoding NIN and Sus genes were isolated. They were 1 932 and 2 418 bp in length， respectively， encoding putative proteins of 643 and 805 amino acid. Sequence alignment results showed that both NIN and Sus were highly conserved， and shared more than 70% sequence homology with other plants from NCBI and almost 100% homology with citrus fruit. GenBank accession number for NIN and Sus from Ougan fruit was KF694988 and KF694989， respectively. Relative molecular mass of the putative NIN and Sus protein was 72.18 and 92.19 ku， respectively， and the isoelectric point （pI） was 6.882 and 6.051， respectively. Phylogenetic analysis showed that putative NIN protein belonged to Group α， which might localize in plastid， and putative Sus protein belongs to Sus Ⅰ. Results of realtime quantitative PCR showed that the mRNA level of both two genes accumulated highly in stem， then leaf， peel and flesh in descending order. The transcription level of NIN in peel was higher than that in flesh， while the transcription level of Sus in peel was relative to that in flesh. At different development stages of fruit， both genes expressed the highest at 120 after full bloom and then maintained at a low level until fruit maturation. The study showed that both NIN and Sus genes expressed with tissuespecificity， and both expressed at a higher level at the immature stage and decreased towards the ripening stage.

Key words: Ougan fruit, neutral invertase, sucrose synthase, gene clone, gene expression