Mutant strain selection

The bacterial and mammalian cell assays for gene mutation were developed to measure statistically significant increases in the numbers of mutant colonies derived from rare events many millions of exposed cells must be plated out to allow the assessment of mutation frequency. The Salmonella typhimurium reverse mutation assay ( Ames test) is carried out in a variety of different mutant strains selected to identify the various classes of mutation. The test generates many hundreds of Petri dishes for counting and is not practical for profiling. 

Books and reviews detailing methods of mutant strain selection... 

Mutation. For industrial appHcations, mutations are induced by x-rays, uv irradiation or chemicals (iiitrosoguanidine, EMS, MMS, etc). Mutant selections based on amino acid or nucleotide base analogue resistance or treatment with Nystatin or 2-deoxyglucose to select auxotrophs or temperature-sensitive mutations are easily carried out. Examples of useful mutants are strains of Candida membranefaciens, which produce L-threonine Hansenu/a anomala, which produces tryptophan or strains of Candida lipolytica that produce citric acid. An auxotrophic mutant of S. cerevisiae that requires leucine for growth has been produced for use in wine fermentations (see also Wine). This yeast produces only minimal quantities of isoamyl alcohol, a fusel oil fraction derived from leucine by the Ehrlich reaction (10,11). A mutant strain of bakers yeast with cold-sensitive metaboHsm shows increased stabiUty and has been marketed in Japan for use in doughs stored in the refrigerator (12). 

Fig. 4. Pleiotropism of Phycomyces strains selected as phototropic defective mutants. Mutants madA and madB are defective in all photoresponses, mutants madD, E, F, G only in tropic and growth responses...

Selection of mutants was done by increasing the chloramphenicol concentration in liquid BG-11. Control and mutant strains were grown at a light intensity of 30 pmol photons-m"2 s at 45°C with air/C02 bubbling. 

Broad spectrum, 4th generation cephalosporin demonstrating a low potential for resistance due to lack of p-lactamase induction and low potential for selection of resistant mutant strains as effective as ceftazidime and cefotaxime in comparative trials twice daily dosing may add economic advantage... 

A large variety of inhibitors of RNA synthesis have been identified. Some of these inhibitors have proved useful in elucidating transcription mechanisms, and some have facilitated selection for superior mutant strains with enzymes that are resistant to their inhibition. The inhibitors fall into three classes (fig. 28.22). 

Today, the yield is even higher than ever by selecting better mutant strains, and improving medium and fermentation techniques. The major steps in penicillin production are as follows (Queener and Swartz, 1979) ... 

Further confirmation that the model developed for 5. cerevisiae is an accurate one for Candida comes from studies of a small set of spontaneous L-733,560-resistant mutants (CAI4R560, NR1-4) and strains selected for resistance by mutagenesis (CA-2 and M-2) (Kurtz et al., 1995, 1996 Douglas et al., 1997). These mutants have enzyme activity that was 5 to 40 times as resistant to inhibition by L-733,560 as the parent strain (Douglas et al., 1997). They are cross-resistant to other glucan... 

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