16S ribosomal RNA
16S ribosomal RNA (or 16S rRNA) is a component of the 30S subunit of prokaryotic ribosomes. It is approximately 1.5kb (or 1500 nucleotides) in length.[2] The genes coding for it are referred to as 16S rDNA and are used in reconstructing phylogenies.
Multiple sequences of 16S rRNA can exist within a single bacterium.[3]
Functions
It has several functions:
- Like the large (23S) ribosomal RNA, it has a structural role, acting as a scaffold defining the positions of the ribosomal protein
- The 3' end contains the anti-Shine-Dalgarno sequence, which binds upstream to the AUG start codon on the mRNA. The 3'-end of 16S RNA binds to the proteins S1 and S21 known to be involved in initation of protein synthesis; RNA-protein cross-linking by A.P. Czernilofsky et al. (FEBS Lett. Vol 58, pp 281–284, 1975).
- Interacts with 23S, aiding in the binding of the two ribosomal subunits (50S+30S)
- Stabilizes correct codon-anticodon pairing in the A site, via a hydrogen bond formation between the N1 atom of Adenine (see image of Purine chemical structure) residues 1492 and 1493 and the 2'OH group of the mRNA backbone
Structure
Universal Primers
The 16SrRNA gene is used for phylogenetic studies[4] as it is highly conserved between different species of bacteria and archaea.[5] Carl Woese pioneered this use of 16S rRNA. In addition to these, mitochondrial and chloroplastic rRNA are also amplified.
Universal (or quasi-universal as it does not pick up some recently discovered hydrothermal archaea species belonging to the phylum Nanoarchaeota[6]) PCR primers are used to amplify the 16SrRNA gene providing the phylogenetic information, the most common universal primer pair was devised by Weisburg et al.[4] and are currently referred to 27F and 1492R, however, for some applications shorter amplicons may be necessary for example for 454 sequencing with Titanium chemistry (500-ish reads are ideal) the primer pair 27F-534R covering V1 to V3.[7]
Sequence analysis of the 16S rRNA sequences is done with the help of several primers, called "universal primers." These primers target the conserved region of 16S rRNA gene and amplify the target in parts. Finally the several amplified parts could be assembled together to have the entire sequence of the complete 16S rRNA. Some of the primers are listed below:
| Primer name |
Sequence (5'-3') |
Reference |
| B27F |
AGA GTT TGA TCC TGG CTC AG |
[8] |
| U1492R |
GGT TAC CTT GTT ACG ACT T |
same as above |
| 928F |
TAA AAC TYA AAK GAA TTG ACG GG |
[9] |
| 336R |
ACT GCT GCS YCC CGT AGG AGT CT |
as above |
| 1100F |
YAA CGA GCG CAA CCC |
|
| 1100R |
GGG TTG CGC TCG TTG |
|
| 337F |
GAC TCC TAC GGG AGG CWG CAG |
|
| 907R |
CCG TCA ATT CCT TTR AGT TT |
|
| 785F |
GGA TTA GAT ACC CTG GTA |
|
| 805R |
GAC TAC CAG GGT ATC TAA TC |
|
| 533F |
GTG CCA GCM GCC GCG GTA A |
|
| 518R |
GTA TTA CCG CGG CTG CTG G |
|
PCR applications
In addition to highly conserved primer binding sites, 16S rRNA gene sequences contain hypervariable regions that can provide species-specific signature sequences useful for bacterial identification. As a result, 16S rRNA gene sequencing has become prevalent in medical microbiology as a rapid, accurate alternative to phenotypic methods of bacterial identification.[10] Although it was originally used to identify bacteria, 16S sequencing was subsequently found to be capable of reclassifying bacteria into completely new species, or even genera.[11][12] It has also been used to describe new species that have never been successfully cultured.[13][14]
References
- ^ Schluenzen F, Tocilj A, Zarivach R, Harms J, Gluehmann M, Janell D, Bashan A, Bartels H, Agmon I, Franceschi F, Yonath A Structure of functionally activated small ribosomal subunit at 3.3 angstroms resolution (2000). "Structure of functionally activated small ribosomal subunit at 3.3 angstroms resolution". Cell 102 (5): 615–23. doi:10.1016/S0092-8674(00)00084-2. PMID 11007480.
- ^ greengenes.lbl.gov - Aligned 16S rDNA data and tools
- ^ Case RJ, Boucher Y, Dahllöf I, Holmström C, Doolittle WF, Kjelleberg S (January 2007). "Use of 16S rRNA and rpoB Genes as Molecular Markers for Microbial Ecology Studies". Appl. Environ. Microbiol. 73 (1): 278–88. doi:10.1128/AEM.01177-06. PMC 1797146. PMID 17071787. http://aem.asm.org/cgi/pmidlookup?view=long&pmid=17071787.
- ^ a b 16S ribosomal DNA amplification for phylogenetic study W G Weisburg, S M Barns, D A Pelletier and D J Lane; J Bacteriol. 1991 January; 173(2): 697-703
- ^ Coenye T, Vandamme P (November 2003). "Intragenomic heterogeneity between multiple 16S ribosomal RNA operons in sequenced bacterial genomes". FEMS Microbiol. Lett. 228 (1): 45–9. doi:10.1016/S0378-1097(03)00717-1. PMID 14612235.
- ^ Huber H, Hohn MJ, Rachel R, Fuchs T, Wimmer VC, Stetter KO (2002). "A new phylum of Archaea represented by a nanosized hyperthermophilic symbiont". Nature 417 (6884): 63–7. doi:10.1038/417063a. PMID 11986665.
- ^ http://www.hmpdacc.org/tools_protocols.php#sequencing
- ^ Universal Bacterial Identification by PCR and DNA Sequencing of 16S rRNA Gene. PCR for Clinical Microbiology, 2010, Part 3, 209-214
- ^ Weidner S, Arnold W, Pühler A (1996). "Diversity of uncultured microorganisms associated with the seagrass Halophila stipulacea estimated by restriction fragment length polymorphism analysis of PCR-amplified 16S rRNA genes". Appl Env Microbiol 62 (3): 766–71. http://aem.asm.org/cgi/reprint/62/3/766.pdf.
- ^ J. E. Clarridge III (2004). "Impact of 16S rRNA gene sequence analysis for identification of bacteria on clinical microbiology and infectious diseases". Clin Microbiol Rev 17 (4): 840–862. doi:10.1128/CMR.17.4.840-862.2004.
- ^ Weisburg WG, Barns SM, Pelletier DA, Lane DJ (1991). "16S ribosomal DNA amplification for phylogenetic study". J Bacteriol 173 (2): 697–703. PMC 207061. PMID 1987160. http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pmcentrez&artid=207061.
- ^ Brett P J, DeShazer D, Woods DE (1998). "Burkholderia thailandensis sp. nov., a Burkholderia pseudomallei-like species". Int J Syst Bacteriol 48: 317–320. doi:10.1099/00207713-48-1-317. PMID 9542103.
- ^ Schmidt TM, Relman DA (1994). "Phylogenetic identification of uncultured pathogens using ribosomal RNA sequences". Methods Enzymol. Methods in Enzymology 235: 205–22. doi:10.1016/0076-6879(94)35142-2. ISBN 9780121821364. PMID 7520119.
- ^ Gray JP, Herwig RP (1996). "Phylogenetic analysis of the bacterial communities in marine sediments". Appl Environ Microbiol 62 (11): 4049–59. PMC 168226. PMID 8899989. http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pmcentrez&artid=168226.
External links
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Mitochondrial
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B bsyn: dna (repl, cycl, reco, repr) · tscr (fact, tcrg, nucl, rnat, rept, ptts) · tltn (risu, pttl, nexn) · dnab, rnab/runp · stru (domn, 1°, 2°, 3°, 4°)
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