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Important Update of the 16S rDNA Primer Evaluation Paper Published in Nucleic Acids Research
Unforntunately for the recommended group M Archaea primer a wrong primer pair was shown. The corresponding paragraph has been corrected. Please update to the latest version of the paper.
Corrections made:
Page 6, left column, line 7-21, the recommended reverse primer S-*-Univ-1392-a-A-15 is wrong and needs to be replaced by S-D-Arch-1041-a-A-18.
The corrected paragraph reads:
In silico evaluation of primer pairs suitable for sequencing technologies like Roche’s 454 (Group M) No archaeal-specific primer pair achieves a full phylum spectrum (Supplementary Table S15). S-D-Arch-0519-a-S-15/S-D-Arch-1041-a-A-18 (A: 76.6%, B: 0.0%, E: 0.0%) shows the best results with respect to a relatively high overall coverage coupled with a high domain specificity. This primer pair covers two out of eight phyla (Crenarchaeota and Euryarchaeota), but the phylum spectrum increases remarkably to six detected phyla if one mismatch is allowed (A: 92.8%, B: 0.0%, E: 0.0%). Detection of the four additional phyla (AAG, Korarchaeota, MHVG I and MHVG II) is likely due to a middle mismatch position in the reverse primer. Amplification of GoC-Arc-109-D0-C1-M0 and Nanoarchaeaota remains challenging due to more than one mismatch. In summary, S-D-Arch-0519-a-S-15/S-D-Arch-1041-a-A-18 is the most suitable primer pair with a 540 bp amplicon spanning HV regions 4–6 and excellent domain specificity. The frequent use of HV region six in diversity analysis makes this pair particularly interesting for comparative analysis (35,44,45).
and on page 7, right column, line 55-59 the reverse primer S-*-Univ-1392-a-A-15 needs to be replaced by S-D-Arch-1041-a-A-18.
The corrected paragraph reads:
For Group M, only 32 sequences of sufficient length were available to re-evaluate the recommended archaeal primer pair S-D-Arch-0519-a-S-15/ S-D-Arch-1041-a-A-18. Thus the Archaea primer pairs were excluded from further validation.
We apologise for these errors and any inconveniences caused.
Paper Summary (unchanged, just for your orientation)
16S ribosomal RNA gene (rDNA) amplicon analysis remains the standard approach for the cultivation-independent investigation of microbial diversity. The accuracy of these analyses depends strongly on the choice of primers. The overall coverage and phylum spectrum of 175 primers and 512 primer pairs were evaluated in silico with respect to the SILVA 16S/18S rDNA non-redundant reference dataset (SSURef 108 NR). Based on this evaluation a selection of ‘best available’ primer pairs for Bacteria and Archaea for three amplicon size classes (100–400, 400–1000, >1000 bp) is provided. The most promising bacterial primer pair (S-D-Bact-0341-b-S-17/S-D-Bact-0785-a-A-21), with an amplicon size of 464 bp, was experimentally evaluated by comparing the taxonomic distribution of the 16S rDNA amplicons with 16S rDNA fragments from directly sequenced metagenomes. The results of this study may be used as a guideline for selecting primer pairs with the best overall coverage and phylum spectrum for specific applications, therefore reducing the bias in PCR-based microbial diversity studies.
Follow this link to read the updated full paper at Nucleic Acids Research
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