Natural isolates usually produce commercially important products in very low concentrations and there fore every attempt is made to increase the productivity of the chosen organism. Increased yields may be achieved by optimizing the culture medium and growth conditions, but this approach will be limited by the organism's maximum ability to synthesize the product. The potential productivity of the organism is controlled by its genome and, therefore, the genome must be modified to increase the potential yield. The cultural requirements of the modified organism would then be examined to provide conditions that would fully exploit the increased potential of the culture, while further attempts are made to beneficially change the genome of the already improved strain. Thus, the process of strain improvement involves the continual genetic modification of the culture, followed by reappraisals of its cultural requirements.
Genetic modification may be achieved by selecting natural variants, by selecting induced mutants and by selecting recombinants. There is a small probability of a genetic change occurring each time a cell divides and when it is considered that a microbial culture will undergo a vast number of such divisions it is not surprising that the culture will become more heterogeneous. The heterogeneity of some cultures can present serious problems of yield degeneration because the variants are usually inferior producers compared with the original culture. However, variants have been isolated which are superior producers and this has been observed frequently in the early stages in the development of a natural product from a newly isolated organism. An explanation of this phenomenon for mycelial organisms may be that most new isolates are probably heterokaryons (contain more than one type of nucleus) and the selection of the progeny of uninucleate spores results in the production of homokaryons (contain only one type of nucleus) which may be superior producers. However, the phenomenon is also observed with unicellular isolates which are certainly not heterokaryons. Therefore, it is worthwhile to periodically plate out the producing culture and screen a proportion of the progeny for productivity; this practice has the added advantage that the operator tends to become familiar with morphological characteristics associated with high productivity and, by selecting 'typical' colonies, a strain subject to yield degeneration may still be used with consistent results.
Therefore, selection of natural variants may result in increased yields but it is not possible to rely on such improvements, and techniques must be employed to increase the chances of improving the culture. These techniques are the isolation of induced mutants and recombination. The most dramatic examples of strain improvement come from the applications of recombinant DNA technology which has resulted in organisms producing compounds which they were not able to produce previously. Furthermore, the advances in these techniques have resulted in very significant improvements in the production of conventional fermentation products. However, it should be remembered that these methods have not replaced mutant isolation but have made an invaluable addition to an impressive repertoire. The techniques of mutant isolation have contributed enormously to the development of present-day industrial strains and these techniques will be considered first. The applications of recombination systems will then be discussed along with the interactions between mutant isolation and recombination in strain improvement advances.
Dulaney and Dulaney (1967) compared the spread in productivity of chlortetracycline of natural variants of Streptomyces viridifaciens with the spread in productivity of the survivors of an ultraviolet treatment. The results of their comparison are shown in Figs 3.3 and 3.4, from which it may be seen that although there are more inferior producers amongst the survivors of the ultraviolet treatment there are also strains producing more than twice the parental level, far greater than the best of the natural variants. The use of ultraviolet light is only one of a large number of physical or chemical agents which increase the mutation-rate — such agents are termed mutagens. The reader is referred to Baltz (1986) and Birge (1988) for accounts of the modes of action of mutagens. The vast majority of induced muta-
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