皱纹盘鲍群体选育第四代成鲍4种多糖裂解酶基因表达的初步研究

Expression of polysaccharidase enzymes in the F_4 adults of a mass selected Pacific abalone strain

根据已发表c DNA序列设计引物扩增了2龄皱纹盘鲍(Haliotis discus hannai Ino)的褐藻酸酶、海带淀粉酶、α-淀粉酶和纤维素酶基因片段并测定其序列,同时用半定量反转录PCR和实时荧光定量PCR技术分析了它们在不同组织中的表达。结果表明:以cDNA为模板的PCR产物序列与已发表的相应基因序列一致;以DNA为模板扩增的a-淀粉酶和纤维素酶基因片段分别含438 bp和667 bp的内含子;在纤维素酶基因片段的外显子和内含子区各检测到2个SNP;4种多糖裂解酶基因均主要在消化腺中表达,褐藻酸酶基因的表达最强,其次为纤维素酶,表明在摄食海带的条件下,褐藻酸是成体皱纹盘鲍可利用的最重要多糖类物质之一。本文的结果为进一步开展皱纹盘鲍"97"选育群体在不同发育期、摄食不同藻类及不同培育环境下的多糖水解酶基因表达研究奠定了基础。

Alginatelyase, laminarinase, α-amylase and cellulase gene segments were amplified in DNA and cDNA templates from the 2-year-old F4 adults of ＇97 selected Pacific abalone strain with primers designed based on published cDNA sequences of these genes. Sequencing of these gene segments revealed one intron of 438 bp in the α-amylase gene, one intron of 667 bp in the cellulase gene, and two SNPS in both exon and intron of the cellulase. Semi-quantitative RT-PCR and Real-time PCR analyses showed that the four enzymes were mainly expressed in the digestive gland, with the alginatelyase gene expression the highest, followed by the cellulose gene, suggesting that alginate may be one of the most important carbohydrate sources in the selected abalone strain fed with kelp. The results in this study may form the basis for further studies on the expression of polysaccharidases of the ＇97 strain at different developmental stages, fed with different macroalgae and under different aquatic environments.