title: Application of 2,4-Dinitrophenylhydrazine (DNPH) in High-Throughput Screening for Microorganism Mutants Accumulating 9α-Hydroxyandrost-4-ene-3,17-dione (9α-OH-AD).
abstract: To develop a quick method for the preliminarily screening of mutant strains that can accumulate 9α-hydroxyandrost-4-ene-3,17-dione (9α-OH-AD), a high-throughput screening method was presented by applying the principle that 2,4-dinitrophenylhydrazine (DNPH) can react with ketones to produce precipitation. The optimal color assay conditions were the substrate androst-4-ene-3,17-dione (AD) concentration at 2.0 g/L, the ratio of AD to DNPH solution at 1:4, and the sulfuric acid and ethanol solution percentages in DNPH solution at 2% and 35%, respectively. This method was used to preliminarily screen the mutants of Rhodococcus rhodochrous DSM43269, from which the three ones obtained could produce more 9α-OH-AD. This DNPH color assay method not only broadens screening methods and increases screening efficiency in microbial mutation breeding but also establishes a good foundation for obtaining strains for industrial application.
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, People's Republic of China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, People's Republic of China.
authors.forename: Fei
authors.initials: F
authors.lastname: Cao
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, People's Republic of China.
authors.forename: Hui
authors.initials: H
authors.lastname: Xiong
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, People's Republic of China.
authors.forename: Yanbing
authors.initials: Y
authors.lastname: Shen
authors.affiliation: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, People's Republic of China.
authors.forename: Min
authors.initials: M
authors.lastname: Wang
mesh_terms: D011232:Chemical Precipitation; D057166:High-Throughput Screening Assays / Q000379:methods*; D007659:Ketones / Q000737:chemistry / Q000378:metabolism; D009154:Mutation; D010088:Oxidoreductases; D010659:Phenylhydrazines / Q000737:chemistry; D012240:Rhodococcus / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*
chemical_list: D007659:Ketones; D010659:Phenylhydrazines; C004787:2,4-dinitrophenylhydrazine; D010088:Oxidoreductases