Radchenko M. Generation of Bacillus subtilis strain-producer with increased accumulation of riboflavin

The thesis is devoted to the obtaining and selection of new riboflavin-producing strains. The aim of the work was to isolate the novel strains of the genus Bacillus from natural sources, namely the surface of vegetable crops, using the fingerprint method and to obtain a producer strain with increased accumulation of riboflavin. In order to search for riboflavin-synthesizing bacteria, 9 samples of potato tubers of the Adretta variety, collected in the fields of Kyiv suburban area, were examined. As a result, 10 cultures with colonies of different colors were isolated from 9 samples of potato tubers. A pure culture of microorganisms with the greatest accumulation of riboflavin was obtained by the method of limiting dilutions for further establishment of its cultural and morphological features. Phylogenetic analysis of the 16S rRNA gene sequence of the producer strain confirmed that the selected strain belongs to the species B. subtilis. The riboflavin-producing strain obtained was included in the «Collection of strains of microorganisms and plant lines for agricultural and industrial biotechnology» of the Institute of Food Biotechnology and Genomics of the National Academy of Sciences of Ukraine under the number IFBG MK-2 and deposited in the «National Depository of Microorganisms» of the D.K. Zabolotny Institute of Microbiology and Virology National Academy of Sciences of Ukraine under ID number IMV B-7797. It was shown that when the strain was cultivated in flasks with a volume of 1 dm3, the accumulation of riboflavin was 13,9 g/dm3 at the 66th hour of cultivation. During the cultivation of strain IFBG MK-1A in a bioreactor with periodic feeding, the accumulation of riboflavin significantly increased (by 65 %) compared to the accumulation of riboflavin in flasks. The strains of B. subtilis IMV B-7797, IFBG MK-1A and IFBG NTG2 are further proposed for use in the industrial technology of riboflavin and to create a recombinant strain based on them for over synthesis of riboflavin.