The 16S rRNA gene is used for phylogenetic studies^[1] as it is highly conserved between different species of bacteria and archaea^[2]. Carl Woese pioneered this use of 16S rRNA^[3]. It is suggested that 16S rRNA gene can be used as a reliable molecular clock because 16S rRNA sequences from distantly related bacterial lineages are shown to have similar functionalities^[4]. Some (hyper)thermophilic archaea (i.e. order Thermoproteales) contain 16S rRNA gene introns that are located in highly conserved regions and can impact the annealing of "universal" primers^[5]. Mitochondrial and chloroplastic rRNA are also amplified.

The most common primer pair was devised by Weisburg et al^[1]. 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^[6]. Often 8F is used rather than 27F. The two primers are almost identical, but 27F has an M instead of a C. AGAGTTTGATCMTGGCTCAG compared with 8F^[7].

Primer name Sequence (5'-3')  Reference

8F  AGA GTT TGA TCC TGG CTC AG [8][9]

U1492R    GGT TAC CTT GTT ACG ACT T     [8][9]

928F    TAA AAC TYA AAK GAA TTG ACG GG   [10]

336R   ACT GCT GCS YCC CGT AGG AGT CT    [10]

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   

27F      AGA GTT TGA TCM TGG CTC AG      [11]

1492R CGG TTA CCT TGT TAC GAC TT  [11]

Reference

[1]     Weisburg WG, Barns SM, Pelletier DA, Lane DJ (January 1991). "16S ribosomal DNA amplification for phylogenetic study". Journal of Bacteriology. 173 (2): 697–703. doi:10.1128/jb.173.2.697-703.1991. PMC 207061. PMID 1987160.

[2]     Coenye T, Vandamme P (November 2003). "Intragenomic heterogeneity between multiple 16S ribosomal RNA operons in sequenced bacterial genomes". FEMS Microbiology Letters. 228 (1): 45–9. doi:10.1016/S0378-1097(03)00717-1. PMID 14612235.

[3]     Woese CR, Fox GE (November 1977). "Phylogenetic structure of the prokaryotic domain: the primary kingdoms". Proceedings of the National Academy of Sciences of the United States of America. 74 (11): 5088–90. Bibcode:1977PNAS...74.5088W. doi:10.1073/pnas.74.11.5088. PMC 432104.

[4]     Tsukuda, Miyuki; Kitahara, Kei; Miyazaki, Kentaro (2017-08-30). "Comparative RNA function analysis reveals high functional similarity between distantly related bacterial 16 S rRNAs". Scientific Reports. 7 (1): 9993. Bibcode:2017NatSR...7.9993T. doi:10.1038/s41598-017-10214-3. ISSN 2045-2322. PMC 5577257. PMID 28855596. Archived from the original on 2018-05-15.

[5]     Jay ZJ, Inskeep WP (July 2015). "The distribution, diversity, and importance of 16S rRNA gene introns in the order Thermoproteales". Biology Direct. 10 (35): 35. doi:10.1186/s13062-015-0065-6. PMC 4496867. PMID 26156036.

[6]     http://www.hmpdacc.org/tools_protocols.php#sequencing Archived 2010-10-30 at the Wayback Machine

[7]     "Primers, 16S ribosomal DNA - François Lutzoni's Lab". lutzonilab.net. Archived from the original on 2012-12-27.

[8]     Eden PA, Schmidt TM, Blakemore RP, Pace NR (April 1991). "Phylogenetic analysis of Aquaspirillum magnetotacticum using polymerase chain reaction-amplified 16S rRNA-specific DNA". International Journal of Systematic Bacteriology. 41 (2): 324–5. doi:10.1099/00207713-41-2-324. PMID 1854644.

[9]     James, Greg (15 May 2018). "Universal Bacterial Identification by PCR and DNA Sequencing of 16S rRNA Gene". PCR for Clinical Microbiology. Springer, Dordrecht. pp. 209–214. doi:10.1007/978-90-481-9039-3_28. ISBN 978-90-481-9038-6.

[10]  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" (PDF). Appl Environ Microbiol. 62 (3): 766–71. PMC 167844. PMID 8975607. Archived (PDF) from the original on 2011-07-15.

[11]  Jiang H, Dong H, Zhang G, Yu B, Chapman LR, Fields MW (June 2006). "Microbial diversity in water and sediment of Lake Chaka, an athalassohaline lake in northwestern China". Applied and Environmental Microbiology. 72 (6): 3832–45. doi:10.1128/AEM.02869-05. PMC 1489620. PMID 16751487.

注意：内容源于维基百科（https://en.wikipedia.org/wiki/16S_ribosomal_RNA#cite_note-HC_Jiang-14），如有错误还请指正。