Study On Influence Of Knockout Of Aspergillus Niger α-glucosidase Genes On Utilization Of Sugar And Production Of Citric Acid

Citric acid is currently the second largest industrial bio-fermentation technology product after ethanol.It is a bulk bio-technology product with significant economic value and strategic significance.It is widely used in food(70%),chemical,pharmaceutical and other industries.At present,about 80%of the production method of citric acid is produced by deep-phase batch fermentation of Aspergillus niger.During the citric acid fermentation process,about 2%of the residual sugar in the fermentation broth cannot be used,and its component is mainly isomaltose.Since isomaltose is mainly catalyzed by α-glucosidase,this study knocked out the α-glucosidase gene in the citric acid production strain,and studied the relationship between the gene knockout and the formation of residual sugar and citric acid production.Using the transformation method of homologous recombination mediated by Agrobacterium tumefaciens,the Aspergillus niger α-glucosidase gene knockout strain KO-49940 and the double knockout strain KO-40261-214233 were constructed,and then the knockout strain and the wild type strain were carried out The comparison of colony morphology,mycelium morphology and spore morphology was observed.The mycelium perimeter and growth curve were measured,and the results showed no significant difference.The detection of sugar and acid content in the fermentation broth of KO-49940,and KO-214233,KO-40261,and KO-40261-214233 by DNS method and HPLC showed that:at the end of 84 h fermentation,the knockout The reducing sugar content in the fermentation broth of the strain and the wild-type strain was basically zero,indicating that the knockout of the glycosidase gene did not significantly affect the rate at which the strain utilizes reducing sugar;but compared with wild-type Aspergillus niger strains,the residual sugar content of the knockout bacteria KO-49940,KO-214233,KO-40261,KO-40261-214233 decreased by about 6.4%,12.9%,12.9%,19.3%,respectively;the content of isomaltose decreased by about 19.1%,23.8%,24.5%and 28.5%,respectively;Citric acid output increased by 1.2%,2.5%,2.5%and 3.7%.It shows that these α-glucosidase gene knockouts are beneficial to reduce the formation of isomaltose,reduce the content of residual sugar,improve the utilization rate of sugar and the yield of citric acid.