Frequency of PHO-constitutive mutants: influence of the culture medium and its buffers

• Grant number: 16/23981-5
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: February 01, 2017
• End date: December 31, 2019
• Field of knowledge: Biological Sciences - Genetics - Molecular Genetics and Genetics of Microorganisms
• Principal Investigator: Beny Spira
• Grantee: Gabriella Trombini Machado
• Host Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

PHO-constitutive mutants are characterized by losing the capacity of regulate the expression of PHO regulon in conditions with excess phosphate. Such mutants are isolated in minimal medium containing glycerol-2-phosphate (G2P) as source of carbon and Tris buffer. The emergence of these mutants follows a typical pattern of adaptive mutants. The actual frequency of PHO constitutive mutants is around 10-7 - 10-8 mutations / cell / generation. Given that PHO-constitutive phenotype derives from the loss of regulatory protein function and these mutations can occur in 6 distinct genes, the frequency of these mutants is very low. The objective of this project is to evaluate the reason for low frequency of PHO-constitutive mutants. One hypothesis is that the emergence of these mutants would be inhibited by toxic effect of the buffer in the culture medium, causing growth inhibition or cell death. To test this possibility, the viability and survival of cells in culture media and the effect of different buffers on the frequency of PHO-constitutive mutants will be analyzed. Also it will be evaluated the dynamics of the emergence of mutants in other systems, such as selection of mutants phoA- and mutants resistant to antibiotics. (AU)