Modern Genomics:How to identify microbial species.
Classic methods dependent upon comparative morphology and culturing fall short for the following reasons -
- Prokaryotes are incredibly diverse with relatively little morphological distinction (can't tell them apart under the microscope).
- Fewer than 1% of the known microbes in any given environment can be cultured (the great plate anomaly).
The Answer - Molecular systemetics based upon 16s rRNA sequences (thanks to Carl Woese, the father of modern microbial systematics).
- The 16S rRNA gene is ideal because much of it is conserved due to structural constraints but some of it is not. This allows for variability in the gene sequence useful in making phylogenetic comparisons.
- Because the gene is only about 1550 bp in length and it portions of it are conserved, it is possible to construct primers that will amplify this gene in a major portion of the organisms present in any given environment.
- After PCR amplification, it is possible to sequence the gene and make comparisons with other members of the microbial community or members of its own species.
Sequencing 16s rDNA from individual organisms has given us the ability map phylogenetic relationships between single celled organisms based upon how similar their 16s genes are. A classic example of this is shown below in the "Tree of Life" published by Norm Pace in 1997.
- Carl Woese - Proc Natl Acad Sci 1990;87 (12): 4576-9Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya.
- Norman Pace - Curr Opin Microbiol. 2005 Dec;8(6):638-42.Phylogenetic diversity and ecology of environmental Archaea.
- Protein translation tool at ExPASy - a tool which allows the translation of a nucleotide (DNA/RNA) sequence to a protein sequence.