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Chapter 86 : Metagenomic Methods for the Identification of Active Microorganisms and Genes in Biodegradation Processes
Metagenomic Methods for the Identification of Active Microorganisms and Genes in Biodegradation Processes, Page 1 of 2< Previous page | Next page > /docserver/preview/fulltext/10.1128/9781555815882/9781555813796_Chap86-1.gif /docserver/preview/fulltext/10.1128/9781555815882/9781555813796_Chap86-2.gif
Isolated microorganisms constitute only a minor fraction of the global microbial diversity, which may comprise millions of species. This chapter discusses some cautionary guidelines that should be followed in the attempts to link phylogeny with function. The simplest and most commonly used approach to identify organisms involved in biodegradation is to isolate microbial strains capable of utilizing the target substrate, xenobiotics, or pollutant as a sole C/N source. Sequence-dependent approaches to identify genes are largely limited by an a priori knowledge of gene(s) mediating the biodegradation processes but have proven invaluable for an understanding of the key enzymes in these metabolic pathways. The majority of control in prokaryotes is thought to occur at the transcriptional level, so the presence of an mRNA is strong evidence for the expression of that gene product. Methods for amplifying genes requiring only one gene-specific primer impose less sequence-dependent bias than standard two-primer PCR amplification procedures. These PCR-based strategies have been used for the recovery of the up- or downstream regions flanking a single PCR primer for the recovery of full-length genes. The minimum number of clones that need to be screened in order to find a gene of interest in the metagenomic library increases with the presence of eukaryotic DNA (a eukaryotic genome is 3 to 140,000 Mbp compared to prokaryotic organisms of 0.6 to 9.5 Mbp). In summary, metagenomic techniques have begun to address both the identity of active organisms and their gene products that mediate biodegradation.
Key Concept Ranking
- Denaturing Gradient Gel Electrophoresis
Classification of microarrays used in environmental studies a