title: Advantage of using inosine at the 3' termini of 16S rRNA gene universal primers for the study of microbial diversity.
abstract: To overcome the shortcomings of universal 16S rRNA gene primers 8F and 907R when studying the diversity of complex microbial communities, the 3' termini of both primers were replaced with inosine. A comparison of the clone libraries derived using both primer sets showed seven bacterial phyla amplified by the altered primer set (8F-I/907R-I) whereas the original set amplified sequences belonging almost exclusively to Proteobacteria (95.8%). Sequences belonging to Firmicutes (42.6%) and Thermotogae (9.3%) were more abundant in a library obtained by using 8F-I/907R-I at a PCR annealing temperature of 54 degrees C, while Proteobacteria sequences were more frequent (62.7%) in a library obtained at 50 degrees C, somewhat resembling the result obtained using the original primer set. The increased diversity revealed by using primers 8F-I/907R-I confirms the usefulness of primers with inosine at the 3' termini in studying the microbial diversity of environmental samples.
authors.affiliation: Department of Biotechnology Engineering, Ben-Gurion University of the Negev, P.O. Box 653, Be'er-Sheva 84105, Israel.
authors.forename: Eitan
authors.initials: E
authors.lastname: Ben-Dov
authors.forename: Orr H
authors.initials: OH
authors.lastname: Shapiro
authors.forename: Nachshon
authors.initials: N
authors.lastname: Siboni
authors.forename: Ariel
authors.initials: A
authors.lastname: Kushmaro
mesh_terms: D001419:Bacteria / Q000145:classification* / Q000235:genetics; D017931:DNA Primers / Q000235:genetics*; D017753:Ecosystem; D020459:Genes, rRNA; D014644:Genetic Variation; D007220:Industrial Waste; D007288:Inosine / Q000235:genetics; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D017422:Sequence Analysis, DNA; D012965:Sodium Chloride; D014871:Water Microbiology
chemical_list: D017931:DNA Primers; D007220:Industrial Waste; D012336:RNA, Ribosomal, 16S; D012965:Sodium Chloride; D007288:Inosine