16S ribosomal RNA
16S ribosomal RNA (or 16S rRNA) is a component of the 30S small subunit of prokaryotic ribosomes. It is 1.542kb (or 1542 nucleotides) in length.[2] The genes coding for it are referred to as 16S rDNA and are used in reconstructing phylogenies, thanks to the work of Carl Woese and George E. Fox.[3]
Multiple sequences of 16S rRNA can exist within a single bacterium.[4]
Contents |
Functions[edit]
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 proteins.
- 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 initiation 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[edit]
Universal Primers[edit]
The 16SrRNA gene is used for phylogenetic studies[5] as it is highly conserved between different species of bacteria and archaea.[6] Carl Woese pioneered this use of 16S rRNA.[3] In addition to these, mitochondrial and chloroplastic rRNA are also amplified.
The most common primer pair was devised by Weisburg et al.[5] and is currently referred to as 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] Often 8F is used rather than 27F. Both primers are almost identical, but 27F has a M (A or C) in stead of a C. AGAGTTTGATCMTGGCTCAG compared with 8F.[8]
| Primer name | Sequence (5'-3') | Reference |
|---|---|---|
| 8F | AGA GTT TGA TCC TGG CTC AG | [9][10] |
| U1492R | GGT TAC CTT GTT ACG ACT T | same as above |
| 928F | TAA AAC TYA AAK GAA TTG ACG GG | [11] |
| 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[edit]
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.[12][13] As a result, 16S rRNA gene sequencing has become prevalent in medical microbiology as a rapid and cheap alternative to phenotypic methods of bacterial identification.[14] 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.[15][16] It has also been used to describe new species that have never been successfully cultured.[17][18]
References[edit]
- ^ 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
- ^ a b Woese, C. R.; G. E. Fox (1977-11-01). "Phylogenetic structure of the prokaryotic domain: The primary kingdoms". Proceedings of the National Academy of Sciences 74 (11): 5088–5090. Bibcode:1977PNAS...74.5088W. doi:10.1073/pnas.74.11.5088. ISSN 0027-8424. PMC 432104. PMID 270744.
- ^ 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.
- ^ a b Weisburg WG, Barns SM, Pelletier DA, Lane DJ (January 1991). "16S ribosomal DNA amplification for phylogenetic study". J Bacteriol. 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): 45–49. doi:10.1016/S0378-1097(03)00717-1. PMID 14612235.
- ^ http://www.hmpdacc.org/tools_protocols.php#sequencing
- ^ http://www.lutzonilab.net/primers/page604.shtml
- ^ Eden PA, Schmidt TM, Blakemore RP, Pace NR (1991). "Phylogenetic Analysis of Aquaspirillum magnetotacticum Using Polymerase Chain Reaction-Amplified 16S rRNA-Specific DNA". Int J Syst Bacteriol. 41 (2): 324–325. PMID 1854644.
- ^ 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.
- ^ Pereira, F.; Carneiro, J.; Matthiesen, R.; van Asch, B.; Pinto, N.; Gusmao, L.; Amorim, A. (4 October 2010). "Identification of species by multiplex analysis of variable-length sequences". Nucleic Acids Research 38 (22): e203–e203. doi:10.1093/nar/gkq865.
- ^ Kolbert, CP; Persing, DH (1999 Jun). "Ribosomal DNA sequencing as a tool for identification of bacterial pathogens.". Current opinion in microbiology 2 (3): 299–305. doi:10.1016/S1369-5274(99)80052-6. PMID 10383862.
- ^ 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.
- ^ 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 978-0-12-182136-4. 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.
External links[edit]
- University of Washington Laboratory Medicine: Molecular Diagnosis | Bacterial Sequencing
- The Ribosomal Database Project
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