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| 1 | Design of 16S rRNA gene primers for 454 pyrosequencing of the human foregut microbiome显示文摘AIM:To design and validate broad-range 16S rRNA primers for use in high throughput sequencing to classify bacteria isolated from the human foregut microbiome.METHODS:A foregut microbiome dataset was constructed using 16S rRNA gene sequences obtained from oral,esophageal,and gastric microbiomes produced by Sanger sequencing in previous studies represented by 219 bacterial species.Candidate primers evaluated were from the European rRNA database.To assess the effect of sequence length on accuracy of classification,16S rRNA genes of various lengths were created by trimming the full length sequences.Sequences spanning various hypervariable regions were selected to simulate the amplicons that would be obtained using possible primer pairs.The sequences were compared with full length 16S rRNA genes for accuracy in taxonomic classification using online software at the Ribosomal Database Project (RDP).The universality of the primer set was evaluated using the RDP 16S rRNA database which is comprised of 433 306 16S rRNA genes,represented by 36 phyla.RESULTS:Truncation to 100 nucleotides(nt)downstream from the position corresponding to base 28 in the Escherichia coli 16S rRNA gene caused misclassification of 87(39.7%)of the 219 sequences,compared with misclassification of only 29(13.2%)sequences with truncation to 350 nt.Among 350-nt sequence reads within various regions of the 16S rRNA gene,the reverse read of an amplicon generated using the 343F/798R primers had the least(8.2%)effect on classification.In comparison,truncation to 900 nt mimicking single pass Sanger reads misclassified 5.0%of the 219 sequences.The 343F/798R amplicon accurately assigned 91.8%of the 219 sequences at the species level.Weighted by abundance of the species in the esophageal dataset,the 343F/798R amplicon yielded similar classification accuracy without a significant loss in species coverage(92%).Modification of the 343F/798R primers to 347F/803R increased their universality among foregut species.Assuming that a typicalpolymerase chain reaction can tolerate 2 mismatches between a primer and a template,the modified 347F and 803R primers should be able to anneal 98%and 99.6%of all 16S rRNA genes in the RDP database.CONCLUSION:347F/803R is the most suitable pair of primers for classification of foregut 16S rRNA genes but also possess universality suitable for analyses of other complex microbiomes. | Carlos W Nossa William E Oberdorf Jφrn A Aas Bruce J Paster Todd Z DeSantis Eoin L Brodie Daniel Malamud Michael A Poles Zhiheng Pei | 2010 | World Journal of Gastroenterology2010，16，33： | 16 |
| 2 | Defining the nor- mal bacterial flora of the oral cavity 显示文摘 | AASJ A PASTER B J STOKES L N | 2005 | Journal of Clinical Mi- crobiology2005，43，11： | 1 |
| 3 | In vitro postanibiotic effect of sparfloxacin and ciprofloxacin against Pseudomonas aeruginosa and Enterococcus faecalis显示文摘 | Paster A Peman J Canton E | 1994 | J Antimicrob Chemother1994，34，： | 1 |
| 4 | Cyclophotocoagulation: A Report by the American Academy of Ophthalmology显示文摘 | Paster S A Singh K Lee D A | 2001 | Am J Ophthal2001，108，11： | 1 |
| 5 | Prevalent bacterial species and novel phylotypes in advanced noma lesions显示文摘 | PASTER B J FALKLER W A ENWONWU C O | 2002 | Journal of Clinical Microbiology2002，40，6： | 1 |
| 6 | In vitro post-anibiotic effect of sparfloxacin and ciprofloxacin against Pseudomonas aeruginosa and Enterococcus faecalis 显示文摘 | Peman J Canton E | 1994 | J Antimicrob Chemother1994，34，5： | 1 |
| 7 | Generalized plasticity and the modelling of soil behavior显示文摘 | Paster M Zienkiewicz O C Chan A H C | 1990 | Int J Numer Anal Meth Geomech1990，14，： | 1 |
| 8 | Emergence of PER-2 and VEB-1a in Acinetobacter baumannii Strains in the Americas显示文摘 | Pasterán F Rapoport M Petroni A | | 0，，： | 1 |
| 9 | Haloanaerobiaceae :A new family of moderately halophilic, obligatory anaerobic bacteria显示文摘 | OREN A PASTER B J WOESE C R | 1984 | Syst Appl Microbiol1984，5，1： | 1 |
| 10 | Role of macrophage inflammatory peptide-2 in cerulein-induced acute pancreatitis and pancreatitis-associated lung injury 显示文摘 | Paster CM Rubbia-Brandt L Hadengue A | 2003 | Lab Invest2003，83，4： | 1 |
| 11 | Particle tracking and the diffusion-reaction equation 显示文摘 | Paster A Bolster D Benson D A | 2013 | Water Resource Research2013，49，： | 1 |
| 12 | Distinct pysiologic properties of mieroglia and blood-home cells in rat brain slices after permanent middle cerebral artery occlusion显示文摘 | Lyons SA Paster A Ohlemeyer C | 2000 | J Cereb Blood Flow Metab2000，20，： | 1 |
| 13 | Prevalent bacterial species and novel phylotypes in advanced Noma Lesions显示文摘 | Paster B J Falkler W A Enwonwu C O | 2002 | Journal of Clinical Microbiology2002，40，6： | 1 |
| 14 | The new VAI-CON^* slag stopper system for oxygen converter steelmaking显示文摘 | Enkner B Paster A Schwelberger J | 2002 | Rev Met Paris2002，，4： | 1 |
| 15 | Defining the normal bacterial flora of the oral cavity 显示文摘 | Asj A Paster B J Stokes LN | 2005 | Journal of Clinical Microbiology2005，43，11： | 1 |
| 16 | Defining the normal bacterial flora of the oral cavity显示文摘 | AAS J A PASTER B J STOKES L N | 2005 | J Clin Microbiol2005，43，11： | 1 |
| 17 | Adenoviral transfer of the melanoma differentiation-associated gene 7 ( mda7 ) induces apoptosis of lung cance cells via up-regulation of the double-stranded RNA-dependent protein kinase(PKR) 显示文摘 | Paster A Vorburger SA Barber GN | 2002 | Cancer Res2002，62，8： | 1 |
| 18 | Role of macrophage inflammatory peptide-2 in cerulein-induced acute pancreatitis and pancreatitis- associated lung injury 显示文摘 | Paster CM Rubbia-Brandt L Hadengue A | 2003 | Lab Invest2003，83，4： | 1 |
| 19 | In vitro postanibiotic effect of sparfloxacin and ciprofloxacin against Pseudomonas aeruginosa and Enterococcus faecalis 显示文摘 | Paster A Peman J Canton E | | J Antimicrob Chemother0，34，5： | 1 |
| 20 | Bacterial diversity in human subgingival plaque显示文摘 | Paster BJ Boches SK Galvin JL Ericson RE Lau CN Levanos VA Sahasrabudhe A Dewhirst FE | 2001 | J Bacteriol2001，183，： | 1 |
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