title: Evaluation of the 23S rRNA gene as target for qPCR based quantification of Frankia in soils.
abstract: The 23S rRNA gene was evaluated as target for the development of Sybr Green-based quantitative PCR (qPCR) for the analysis of nitrogen-fixing members of the genus Frankia or subgroups of these in soil. A qPCR with a primer combination targeting all nitrogen-fixing frankiae (clusters 1, 2 and 3) resulted in numbers similar to those obtained with a previously developed qPCR using nifH gene sequences, both with respect to introduced and indigenous Frankia populations. Primer combinations more specifically targeting three subgroups of the Alnus host infection group (cluster 1) or members of the Elaeagnus host infection group (cluster 3) were specific for introduced strains of the target group, with numbers corresponding to those obtained by quantification of nitrogen-fixing frankiae with both the 23S rRNA and nifH genes as target. Method verification on indigenous Frankia populations in soils, i.e. in depth profiles from four sites at an Alnus glutinosa stand, revealed declining numbers in the depth profiles, with similar abundance of all nitrogen-fixing frankiae independent of 23S rRNA or nifH gene targets, and corresponding numbers of one group of frankiae of the Alnus host infection only, with no detections of frankiae representing the Elaeagnus, Casuarina, or a second subgroup of the Alnus host infection groups.
authors: Texas State University, Department of Biology, 601 University Drive, San Marcos, TX 78666, USA.
authors: Suvidha
authors: S
authors: Samant
authors: Max Planck Institute for Marine Microbiology, Celsiusstrasse 1, D-28359 Bremen, Germany.
authors: Rudolf I
authors: RI
authors: Amann
authors: Texas State University, Department of Biology, 601 University Drive, San Marcos, TX 78666, USA. Electronic address: dh49@txstate.edu.
authors: Dittmar
authors: D
authors: Hahn
mesh_terms: D001431:Bacteriological Techniques / Q000379:methods*; D017931:DNA Primers; D040161:Frankia / Q000235:genetics / Q000302:isolation & purification*; D010088:Oxidoreductases / Q000235:genetics; D012338:RNA, Ribosomal, 23S / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012988:Soil Microbiology
chemical_list: D017931:DNA Primers; D012338:RNA, Ribosomal, 23S; D010088:Oxidoreductases; C029130:nitrogenase reductase
keywords: 23S rRNA; Alnus; Frankia; Nitrogenase; Quantitative PCR; Root nodules; nifH