Genetically engineered Streptomyces: Their construction, effects on carbon mineralization, and survival in a natural soil ecosystem.

摘要

Genetically engineered microorganisms are constructed by techniques in which a DNA fragment from one microorganism is introduced into another using a recombinant DNA vector such as a plasmid or bacteriophage. These genetically engineered microorganisms (GEMs) have potential uses in many fields including agriculture, the food industry, the pharmaceutical industry, in pollution control, and in energy production, among others. Relatively, few studies of environmental release of GEMs have been published, even though the ecological consequences of releasing GEMs into the environment need to be evaluated on a case by case basis. There have been very few reports of ecological effects resulting from release of GEMs into the environment. Streptomyces play an important role in the biodegradation of lignin. During the degradation process Streptomyces excrete lignin peroxidase, a key enzyme involved in degrading the lignin polymer. Recombinant Streptomyces that overproduce this enzyme have been constructed in our laboratory. The risks to the environment of releasing such Streptomyces into soil ecosystems are unknown. Presented in this dissertation are three scientific papers which address this question. The first paper describes the survival and effects of wild-type, mutant and recombinant Streptomyces in a soil ecosystem. Some recombinant strains survived, as well as their wild-types, in a laboratory soil microcosm. One genetic variant, Streptomyces lividans TK23-3651, which overproduced extracellular peroxidase and H{dollar}sb2{dollar}O{dollar}sb2{dollar} was observed to significantly enhance carbon mineralization rates in soil. However, it was unstable both in soil and in laboratory medium. Thus, its long term potential for affecting the environment was considered minimal. The second paper describes the construction of a very stable Streptomyces recombinant, S. lividans TK64.1 which expressed a plasmid pIJ702-encoded lignin peroxidase gene from S. viridosporus genomic DNA and overproduced extracellular lignin peroxidase in laboratory growth media. The third manuscript describes a study of the in vitro and in vivo effects of lignin peroxidase and H{dollar}sb2{dollar}O{dollar}sb2{dollar} on carbon mineralization rates in a soil ecosystem. Recombinants Streptomyces lividans strains TK23.1, TK24.1 and TK64.1, all of which overexpress pIJ702-encoded lignin peroxidase, were found to significantly increase CO{dollar}sb2{dollar} evolution rates in soil in comparison to their wild-types.