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Titre du document / Document title

PCR-induced sequence artifacts and bias : Insights from comparison of two 16S rRNA clone libraries constructed from the same sample

Auteur(s) / Author(s)

ACINAS Silvia G. (1) ; SARMA-RUPAVTARM Ramahi (1) ; KLEPAC-CERAJ Vanja (1) ; POLZ Martin F. (1) ;

Affiliation(s) du ou des auteurs / Author(s) Affiliation(s)

(1) Department of Civil and Environmental Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, ETATS-UNIS

Résumé / Abstract

The contribution of PCR artifacts to 16S rRNA gene sequence diversity from a complex bacterioplankton sample was estimated. Taq DNA polymerase errors were found to be the dominant sequence artifact but could be constrained by clustering the sequences into 99% sequence similarity groups. Other artifacts (chimeras and heteroduplex molecules) were significantly reduced by employing modified amplification protocols. Surprisingly, no skew in sequence types was detected in the two libraries constructed from PCR products amplified for different numbers of cycles. Recommendations for modification of amplification protocols and for reporting diversity estimates at 99% sequence similarity as a standard are given.

Revue / Journal Title

Applied and environmental microbiology    ISSN  0099-2240   CODEN AEMIDF 

Source / Source

2005, vol. 71, no12, pp. 8966-8969 [4 page(s) (article)] (38 ref.)

Langue / Language

Anglais

Editeur / Publisher

American Society for Microbiology, Washington, DC, ETATS-UNIS  (1976) (Revue)

Mots-clés anglais / English Keywords

Bias

;

Bacterioplankton

;

Ribosomal RNA

;

16S-RNA

;

Comparative study

;

Artefact

;

Polymerase chain reaction

;

Mots-clés français / French Keywords

Erreur systématique

;

Bactérioplancton

;

RNA ribosomique

;

RNA 16S

;

Etude comparative

;

Artefact

;

Réaction chaîne polymérase

;

Mots-clés espagnols / Spanish Keywords

Error sistemático

;

Bacterioplanctón

;

RNA ribosómico

;

ARN 16S

;

Estudio comparativo

;

Artefacto

;

Reacción cadena polimerasa

;

Localisation / Location

INIST-CNRS, Cote INIST : 7195, 35400013451449.1670

Nº notice refdoc (ud4) : 17317924



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