关键词: 长足大竹象')">长足大竹象, 内切葡聚糖酶, 基因表达, 酶活性, 正交优化

Abstract: 【Aim】 This study aims to investigate the optimal reaction conditions of endoglucanases from Cyrtotrachelus buqueti, and to explore the key genes of endoglucanases in the gut of C. buqueti. 【Methods】 Single factor and orthogonal optimization experiments were carried out. The optimal reaction conditions for endoglucanases were determined by using 3,5-dinitrosalicylic acid (DNS) method with sodium carboxymethyl cellulose (MC) as the substrate. The endoglucanase-encoding gene sequences in the transcriptomes of different developmental stages of C. buqueti were subjected to bioinformatics analysis. Through correlation analysis between gene expression level and enzyme activity, key endoglucanase genes in different developmental stages of C. buqueti were screened. And the expression levels of key genes of endoglucanases in the gut of C. buqueti of different developmental stages were validated by using qRT-PCR. 【Results】 The results showed that the optimum reaction conditions for endoglucanases in C. buqueti adults were 45℃, pH 5.6, and 2% substrate concentration, with the enzyme activity of 59.85 and 52.87 U/mg in female and male, respectively, while the optimum reaction conditions for endoglucanases in C. buqueti larvae were 35℃, pH 4.8, and 2% substrate concentration, with the enzyme activity of 38.34 U/mg. Moreover, two key endoglucanase genes, c64192_g1 and c57057_g1, were identified, and they exhibited higher expression level in adults than in larvae. 【Conclusion】 The endoglucanase activities in female and male adults of C. buqueti are higher than that in larvae, and two key genes, c64192_g1 and c57057_g1, affecting the endoglucanase activity in C. buqueti were screened. These foundings enrich the endoglucanase source and provide data reference for the heterologous expression of endoglucanase gene in C. buqueti, laying a foundation for the pretreatment of lignocellulose and the development and utilization of biomass energy.

Key words: Cyrtotrachelus buqueti')">Cyrtotrachelus buqueti, endoglucanase, gene expression, enzyme activity, orthogonal optimization