Primers used for qPCR assays, annealing temperatures, target regions, and the specificity of the amplicons cloned from qPCR assays with soil DNAa

Target groupForward primerReverse primerApproximate amplicon length (bp)Annealing temp (°C)% of soil clones belonging to the target groupc
All BacteriaEub338Eub51820053100
Actinobacteria Actino235Eub5183006060
Firmicutes Lgc353Eub51818060100
Bacteroidetes Cfb319Eub51822065100
Acidobacteria Acid31Eub51850050100
All Fungi5.8sITS1f300b53100
Basidiomycota ITS4b5.8sr500b55100
  • a The bacterial qPCR assays target 16S rRNA genes, while the fungal assays target the internal transcribed spacer region found in rRNA genes.

  • b The length of the targeted ITS region can vary significantly between different fungal strains.

  • c The percentage includes only those clones that were nonchimeric and exceeded the 80% confidence threshold for taxonomic assignment using the RDP classifier program.