Construction of Bem46 knockout strain and the role of the Bem46 gene in Aspergillus fumigatus / 中国人兽共患病学报

ABSTRACT

The aims of this study are to construct the Bem46 knockout strain of Aspergillus fumigatus,investigate the effect of the Bem46 in the growth and germination,and observe the sensitivity of the oxidative stress,the osmotic stress,the temperature and the response to caspofungin and nikkomycin.The Bem46 gene was identified by sequence alignment in A.fumigatus genome database.The Bem46 knockout strain was construction by the protoplast.The growth was observed and compared between Ku80 and △Bem46 on different culture medium,such as GMM,GMM containing hydrogen peroxide,formamide,sodium chloride and sorbitol.The growth diameter was measured under the different temperatures.The germination rate was observed and compared by microscope.Results showed that the Bem46 gene,Afu7g04660,contained 1 116 bp bases pair and encoding 311 amino acids.Six transformants were obtained by gene cloning and protoplast method and only one was confirmed by PCR and Southernblot.The △Bem46 grew significantly faster than the control strain on the medium containing sorbitol.And there were no visible difference between △Bem46 and Ku80 on the other medium.The germination rate of the △Bem46 was more retarded than the Ku80 in GMM liquid medium.In conclusion,the Bem46 gene plays a role in osmotic stress in A.fumigatus and involved in osmotic pressure induced by sorbitol.And there is no visible effect in oxidative stress.The Bem46 gene has a positive effect on spore germination.