16S ribosomal RNA
16S rRNA (ribosomal RNA) when translated is the small subunit of the ribosome in Prokaryotes. The majority of research on the microbial populations of biomining microorganisms has focused on the analysis of the 16S rRNA gene.
Determining the 16S rRNA gene of a microorganism has become a routine part of phenotypic characterisation and has replaced many culture-based techniques. As the ribosome is essential to cellular function the gene remains highly conserved and can be used to determine the evolutionary relatedness of microorganisms. The vast range of 16S rRNA gene sequences in online databases makes comparative analysis viable.
The gene that codes for 16S rRNA is ~1550bp containing highly conserved regions interspersed with variable regions. The length of the gene allows for meaningful analysis on variations with in the gene sequence, while being short enough to amplify the whole gene by PCR with ease. Conserved regions of the gene can be used to design universal primers for the amplification of a wide range of microorganisms and variable regions can be used to distinguish various species from one another.
Downsides of using the 16S rRNA gene for population studies:
- Quantifying the 16S rRNA gene can only reveal the abundance of a microorganism in the environment.
- Differences in primer binding affinity can cause bias in PCR, resulting in an overrepresentation of some microbial species.
- Microorganisms often contain more than one copy of the 16S rRNA gene, if this is not taken into account during analysis there will be an overrepresentation of species containing more than one 16S rRNA gene.
- Heterogeneity (up to 5%) in 16S rRNA genes within the same organism has been reported (Amann et al. 2000; Mylvaganam & Dennis 1992).
Amann, G, Stetter, KO, Llobet-Brossa, E, Amann, R & Anton, J 2000, 'Direct proof for the presence and expression of two 5% different 16S rRNA genes in individual cells of Haloarcula marismortui', Extremophiles, vol. V4, no. 6, pp. 373-6.
Mylvaganam, S & Dennis, PP 1992, 'Sequence heterogeneity between the two genes encoding 16S rRNA from the halophilic archaebacterium Haloarcula marismortui', Genetics, vol. 130, no. 3, pp. 399-410.