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Information × Registration Number 0222U000895, 0120U104114 , R & D reports Title Creation of yeast producers of new flavin antibiotics popup.stage_title Head Dmytruk Konstantyn V., Доктор біологічних наук Registration Date 21-01-2022 Organization Institute of Cell Biology of the National Academy of Sciences of Ukraine popup.description2  Report: 86 pages, 47 figures, 54 sources. Object of research: the process of biosynthesis of aminoriboflavin and roseoflavin from riboflavin and flavin mononucleotide. The purpose of the study: construction of recombinant strains of yeast Candida famata, capable of overproduction of antibiotics aminoriboflavin and roseoflavin. Research methods: genetic engineering, microbiological, biochemical, bioinformatics analysis. A system of multicopy integration of target DNA sequences into the genome of yeast Candida famata has been developed, CRISPR-Cas9 system for C. famata has been optimized, rosB, rosA and rosC genes of S. davaonensis were designed, plasmids for multicopy integration of rosB, rosA and rosC genes under the control of a strong promoter has been constructed. The constructed plasmids were used for selection of C. famata yeast strains with enhanced expression of the corresponding genes. An aminoriboflavin producer based on the yeast C. famata able to oversynthesis of FMN by overexpression of rosB gene has been constructed. Chromatographic and mass spectrometric analysis revealed aminoriboflavin, which accumulated recombinant strain in culture medium at a concentration of 5.2 mg/l. The Pichia pastoris strain with increased expression of FMN1, rosB, rosA and rosC genes was constructed. The engineered strain produced 3 mg/l of roseoflavin. The scheme of isolation and purification of aminoriboflavin and roseoflavin from the culture medium of the constructed strains was developed and preparative amounts of these antibiotics were obtained. Cultivation conditions were optimized to maximize the accumulation of these antibiotics by the constructed strains. The conditions for culturing the roseoflavin producer in a bioreactor have been optimized. The recombinant P. pastoris strain produced 46 mg/L of roseoflavin during cultivation in a 1 L laboratory bioreactor. Key words: genetic engineering, yeast, Candida famata, Pichia pastoris, roseoflavin, aminoriboflavin, Strep Product Description popup.authors Andreeva Yulia Andrijivna Dmytruk Olena Valerijivna Petrovska Yana Oleksandrivna. Sibirny Andrij Andrijovych Fayura Lubov Romanivna Fedorovych Dariya Vasylivna popup.nrat_date 2022-03-09 Close