Method of constructing a recombinant Bacillus subtilis that can produce specific-molecular-weight hyaluronic acids

A method for making a Bacillus subtilis bacterium cell culture capable of providing specific molecular weight hyaluronic acid, said method comprising the steps of:

• 1) transforming said bacterium with a hasA gene which encodes a hyaluronan synthase, said gene is integrated into the chromosome of said bacterium by use of plasmid pAX01:
• 2) transforming said bacterium with genes tuaD which encodes an UDP-glucose dehydrogenase, glmU which encodes a UDP-N-acetylglucosamine pyrophosphorylase, gtaB which encodes a UDP-glucose pyrophosphorylase, glmM which encodes a mutase, and glmS which encodes an amino transferase, said genes are connected in series and inserted into vector pP43NMK, said vector is transformed into the bacterium;
• 3) coexpressing in said bacterium a hyaluronidase gene fused with a regulatory DNA fragment containing a promoter, a ribosome binding site (RBS) sequence and encoding a signal peptide, said DNA fragment is integrated onto the chromosome of the bacterium;
• wherein said bacterium expresses hyaluronic acid that is acted on by the hyaluronidase to provide specific molecular weight hyaluronic acid in said cell culture.