银耳芽孢的诱导培养及其差异基因片段的克隆

Induction Culture and Isolation Inductive Differentially Expressed Gene Fragments from Yeast-like Conidia of Tremella fuciformis

摘要:

为了探讨银耳芽孢内源诱导型启动子克隆的有效方法，克隆银耳芽孢诱导型差异基因片段，本研究采用乙醇和纤维二糖两种碳源对银耳芽孢进行诱导培养，对其生长情况进行了比较分析，并用cDNA-AFLP技术对差异培养的细胞进行差异表达基因克隆及分析。结果表明乙醇、纤维二糖诱导碳源培养银耳芽孢与葡萄糖相比具有很大差异，其中乙醇培养细胞脱氢酶类活性提高168%，纤维二糖培养细胞最大生长量减小32.2%。从两种差异培养的细胞中克隆得到14条特异性的差异表达基因片段，其中5条在乙醇、纤维二糖样品中各有表达的差异序列，与编码细胞色素C氧化酶、细胞色素b蛋白的基因序列同源，该基因的表达与乙醇代谢途径及胁迫诱导相关。银耳芽孢诱导培养及其诱导差异基因片段的克隆是建立食用菌诱导型表达系统一次有益尝试，为构建高效银耳生物反应器提供一定的理论参考。

Abstract:

In order to probe an effective way of cloning the endogenous inductive promoter and isolating inductive differentially-expressed gene fragments from Yeast-like conidia of Tremella fuciformis, conidia was cultured by two carbon source inducers(ethanol and cellobiose). The inductive differentially-expressed gene fragments were isolated by cDNA-AFLP from the two kinds of differential cells after the growth status analysis. Result showed that, the dehydrogenase activity of the cells cultured with ethanol was 168% higher than that with glucose. The maximum growth rate of the cells cultured with cellobiose reduced by 32.2%. 14 differentially-expressed gene fragments were isolated from the tow differential cells 5 of these gene fragments expressed separately in the cells cultivated with ethanol and cellobiose, which were homologous with the sequence Codes of cytochrome C oxidase and cytochrome b. The gene expression was related to ethanol metabolic and stress induction. This research mayb be of benefit to construction of edible fungus inducible expression system and highly effective bioreactor of Tremella fuciformis.