Identification And Function Analysis Of Aspergillus Niger Hyperosmotic Pressure Regulating Promoter

Aspergillus niger,as a food-grade safe filamentous fungus,has an efficient protein secretion system and is widely used in the expression of homologous and heterologous proteins.Therefore,it is widely used in food,cosmetics,medicine and other fields as production strains.Promoter plays a key role in eukaryotic protein expression.promoter regulates transcription process initiation and downstream gene expression in aspergillus niger.In A.niger,it is very important to have an efficient promoterimprove in order to getting more expression of the target product with.A.niger CGMCC10142 was used as the starting strain in this paper,the promoter probe and gene recombination technique were used to screen the ideal promoter in this strain,and analyze the filtered promoter function.The specific experimental methods are as follows:It constructed pn3 vector as a probe carrier of A.niger promote,which with gene rfp and HYG.By means of electrical transformation,pN3 plasmids were put into Agrobacterium tumefaciens,and then agrobacterium infectingA,niger.the nucleotide sequence with priming activity was obtained in strains that detected reporter gene expression.BLAST analysis of cloned nucleotide sequence with NCBI published genome sequence CBS 513.88 A.niger.As a result,the sequence was 100%similar to the upstream base sequence of the glycoside hydrolase gene,identified as the Pgh promoter.Based on the promoter prediction website promoter 2.0 protect the promoter screened in this paper is predicted.The results show that this promoter may have priming activity.The results showed that the promoter had universal promoter TATA frame and caat-box,and A.niger common sequence.Five A.niger strains with different PgH promoter lengths were analysised.Six strains were named M-pg1,M-pg2,M-pg3,M-pg4,M-pg5,M-pg6.Strain growth and reporter gene expression were observed on the medium with different concentrations of hygromycin and glucose.Finally,it was found that the promoter PgH4 had higher priming activity only under the culture condition of high,and it could be induced by glucose.Almost no startup function is displayed when the promoter is shortened to PgH5.Finally,A.niger was screened for PgH promoter with good priming activity,and found that PgH4 promoter is an inducible promoter with glucose-induced properties.It can be used for the expression of the target gene of A.niger on a higher concentration of glucose medium.