MOLECULAR BREEDING OF A BIOTIN-HYPERPRODUCING SERRATIA-MARCESCENS STRAIN

We previously reported that an acidomycin-resistant mutant of Serratia marcescens Sr41, SB304, and a mutant that was derived from SB304 and was resistant to a higher concentration of acidomycin, SB412, produced 5 and 20 Mg Of D-biotin, respectively, per liter of a medium containi ng sucrose and urea (N. Sakurai, Y. Imai, M. Masuda, S. Komatsubara, a nd T. Tosa, Appl. Environ. Microbiol. 59:2857-2863, 1993). In order to increase the productivity of D-biotin, the biotin (bio) operons were cloned from strains SB412, SB304, and 8000 (wild-type strain), and pLG M412, pLGM304, and pLGW101, respectively, were obtained through subclo ning. These plasmids harbored 7.2-kb DNA fragments coding for the bioA BFCD genes on a low-copy-number vector and were introduced into SB304, SB412, and 8000. Among the resulting recombinant strains, SB412(pLGM3 04) exhibited the highest D-biotin production (200 mg/liter) in the pr oduction medium. The plasmid was stably maintained in cells. Unexpecte dly, SB412(pLGM412) grew very slowly, and the D-biotin productivity of this recombinant strain was not evaluated because pLGM412 was unstabl e.