Abstract:To investigate the potential functions of Unigene0004454 gene, which encodes a hydrophobin in Ganoderma lucidum ZJ-1 strain, this study adopted approaches involving gene cloning, bioinformatics analysis, promoter element analysis, and heterologous expression experiment. The results indicated that the full length of Unigene0004454 gene was 436 bp, a cDNA length 330 bp, and encoded 109 amino acids. Amino acid sequence analysis demonstrated that it has the typical cysteine structure characteristic of hydrophobins, and showed a closer evolutionary relationship with hydrophobins from Ganoderma leucocontextum and Ganoderma sinense. The promoter region of Unigene0004454 contains core promoter element (such as TATA-BOX), multiple hormone responsive elements (such as ABRE), and environmental stress inducible elements (such as MBS). It is speculated that the physiological function of Unigene0004454 may be related to the growth and development of the strain. The recombinant plasmid pET-32a-Unigene0004454 was constructed and transferred into E. coli BL21(DE3) competent cells for expression, followed by SDS-PAGE and Western blotting validation. The results indicated that the recombinant protein with a molecular weight of approximately 27 kDa was successfully expressed in BL21 strain, thereby verifying the feasibility of heterologous expression for Unigene0004454. This study lays a foundation for further elucidation of the physiological functions of Unigene0004454 gene and the development of hydrophobin-based applications.
