Question

Please answer this two part question;

Explain the biochemical mechanism behind heat- and cold-sensitive mutations. What type of mutations would typically lead to a heat-sensitive phenotype and what type of mutations would typically lead to a cold-sensitive phenotype. Also, please describe how auxotrophic mutants are isolated

Answer 1

A.

Most temperature-sensitive mutations affect proteins, and cause loss of protein function at the non-permissive temperature. The permissive temperature is one at which the protein typically can fold properly, or remain properly folded. At higher temperatures, the protein is unstable and ceases to function properly. These mutations are usually recessive in diploid organisms.

  The permissive temperature is the temperature at which a temperature-sensitive mutant gene product takes on a normal, functional phenotype.[When a temperature-sensitive mutant is grown in a permissive condition, the mutated gene product behaves normally (meaning that the phenotype is not observed), even if there is a mutant allele present. This results in the survival of the cell or organism, as if it were a wild type strain. In contrast, the nonpermissive temperature or restrictive temperature is the temperature at which the mutant phenotype is observed.

B. A method for the selection of auxotrophic mutants in Aspergillus nidulans using N-glycosyl-polifungin has been developed. The method results in about 9000- fold enrichment. All types of auxotrophs were obtained. Under certain conditions preferential selection of nucleic acid hydrolysate-requiring auxotrophs was observed. The frequency of occurrence of these mutants was about 50 % of all colonies tested. Cysteine auxotrophs of A. nidulans were obtained for the first time by means of this method.