Fig 1.
dsrB gBlocks and their primer-binding sites.
The 12 gBlocks were selected to cover the currently known phylogenetic sequence diversity of dsrAB genes from sulfate-reducing bacteria and archaea. The five dsrAB gene phylogenetic superclusters are abbreviated by capital letters in brackets, following each numbered gBlock D: Deltaproteobacteria. E: Environmental cluster I. F: Firmicutes FACA group sensu lato. N: Nitrospirae. A: Archaeoglobus. Dark blue circle: gBlocks that contain the primer-binding site of DSRp2060F/DSR4R. Green circle: gBlocks with the primer binding site of DSR1728Fmix/DSR4Rmix. Red circle: gBlocks that contain the primer-binding sites of DSR1762Fmix/DSR2107Rmix. Some gBlocks include primer-binding sites for two primer sets. They have been placed in the overlapping areas of the respective primer set circles indicated by teal, purple, and mixed yellow colors. The central white area in the middle of the Venn diagram contains four gBlocks with primer-binding sites for all three primer sets used in this study.
Fig 2.
Theoretical amplification performance of all gBTMs by different qPCR assays.
(A) Theoretical template amplification percentages of each gBTMs by different qPCR assays. (B) Comparison of theoretical template amplification percentages of each gBTMs among different experimental conditions. Gray (Comparison 1): ratio of theoretical template amplification percentages of each gBTMs by the DSR1728Fmix/DSR4Rmix assay to those by the DSRp2060F/DSR4R assay at each experimental condition. Black (Comparison 2): ratio of theoretical template amplification percentages of each gBTMs by the DSR1762Fmix/DSR2107Rmix to those by the DSRp2060F/DSR4R assay at each experimental condition. White (Comparison 3): ratio of theoretical template amplification percentages of each gBTMs by the DSR1762Fmix/DSR2107Rmix to those by the DSR1728Fmix/DSR4Rmix assay at each experimental condition. The degeneracy of DSRp2060F/DSR4R, DSR1728Fmix/DSR4Rmix assay, and DSR1762Fmix/DSR2107Rmix are 4/1, 77/10, and 98/29, respectively.
Fig 3.
Comparison of theoretical template amplification percentages and empirical qPCR quantification results.
(A) The ratio of theoretical template amplification percentages to empirical qPCR quantification results under high template concentration conditions. (B) The ratio of theoretical template amplification percentages to empirical qPCR quantification results under a low template concentration. Gray (Comparison 1): Comparison between theoretical template amplification percentages and empirical qPCR quantification results of the DSRp2060F/DSR4R assay under different conditions. Black (Comparison 2): Comparison between theoretical template amplification percentages and empirical qPCR quantification results of the DSR1728Fmix/DSR4Rmix assay under other conditions. White (Comparison 3): Comparison between theoretical template amplification percentages and empirical qPCR quantification results of the DSR1762Fmix/DSR2107Rmix assay under different conditions. The degeneracy of DSRp2060F/DSR4R, DSR1728Fmix/DSR4Rmix assay, and DSR1762Fmix/DSR2107Rmix are 4/1, 77/10, and 98/29, respectively.
Fig 4.
Pairwise comparison of empirical qPCR results between different qPCR assays under high (A) and low (B) template concentration conditions.
Gray (Comparison 1): Comparison of empirical qPCR results between the DSR1728Fmix/DSR4Rmix assay and the DSRp2060F/DSR4R assay. Black (Comparison 2): Comparison of empirical qPCR results between the DSR1762Fmix/DSR2107Rmix assay and the DSRp2060F/DSR4R assay. White (Comparison 3): Comparison of empirical qPCR results between the DSR1762Fmix/DSR2107Rmix assay and the DSR1728Fmix/DSR4Rmix assay. The degeneracy of DSRp2060F/DSR4R, DSR1728Fmix/DSR4Rmix assay, and DSR1762Fmix/DSR2107Rmix are 4/1, 77/10, and 98/29, respectively.