Genetically modified clostridial, sucrose perbutyl acetonicam type microorganism with genome sequence specific conversion, its manufacturing method, and use thereof

摘要

PROBLEM TO BE SOLVED: To provide: novel high butanol-producing bacteria, which have excellent safety and are expected to reduce facility costs and waste disposal costs by causing Clostridium saccharoperbutylacetonicum microorganisms to delete functions of enzymes thereof for production of by-products, such as acetic acid and butyric acid, without depending on insertion of foreign genes; and a method for producing the same.SOLUTION: The present invention provides a Clostridium saccharoperbutylacetonicum microorganism, which lacks the function of butyric acid-producing enzyme gene involved in the pathway of producing butyric acid from butyryl CoA, where the gene does not include any insertion and/or deletion. Also provided is a method for producing a microorganism deficient in butyric acid-producing enzyme gene. The method comprises: introducing both a DNA encoding a chimeric RNA consisting of crRNA and tracrRNA, which contains a sequence complementary to the target strand of the target nucleotide sequence in the butyric acid-producing gene, and a DNA encoding a fusion protein of mutant Cas protein and deaminase, which has inactivated cleavage ability of at least one DNA strand, into a Clostridium saccharoperbutylacetonicum microorganism using a vector capable of reproducing in the microorganism to substitute a specific base within the target nucleotide sequence to cause the microorganism to loss the function of the enzyme gene. Further provided is a method for producing butanol by culturing the microorganism deficient in butyric acid-producing enzyme gene.SELECTED DRAWING: None