999 Pectobacterium atrosepticum 90% >0 999 Photorhabdus asymbioti

999 Pectobacterium atrosepticum 90% >0.999 Photorhabdus asymbiotica 96% >0.999 Plesiomonas shigelloides 93% >0.999 Pragia fontium 100% >0.998 Proteus mirabilis 98% >0.999 Providencia rustigianii 93% >0.999 Rahnella aquatilis 92% >0.999 Raoultella ornithinolytica 94% >0.999 Salmonella enterica 101% >0.999 Salmonella enterica subsp. enterica serovar Selleckchem RG-7388 gallinarum 95% >0.998 Serratia liquefaciens

94% >0.999 Shigella dysenteriae 98% >0.999 Tatumella ptyseos 101% >0.999 Trabulsiella guamensis 95% >0.999 Yokenella regensburgei 96% >0.999 Yersinia enterocolitica 98% >0.999 Campylobacter jejuni 89% >0.999 Vibrio cholerae 85% >0.996 Borrelia burgdorferi 90% >0.999 Treponema denticola 82% >0.999 *No 16 S rRNA gene sequence available in the Ribosomal Database Project. Laboratory quantitative assay validation

using pure plasmid standards and mixed templates Assay quantitative validation For the assay quantitative validation, we followed the Minimum Information for publication of Quantitative real-time PCR Experiments, or the MIQE guidelines whenever applicable [10]. The MIQE guidelines were complemented with additional tests to determine assay performance in the presence of background fungal and human genomic DNA. In our experimental design, we included seven template conditions: plasmid standards alone and plasmid standards with 0.5 ng C. albicans genomic DNA (ATCC) and with 0.5 ng, 1 ng, 5 ng, and 10 ng of human genomic DNA per reaction in 10 μl reactions and plasmid standards Cell press alone in 5 μl reactions. For each condition www.selleckchem.com/products/pf-03084014-pf-3084014.html assessed, three qPCR runs were performed to assess reproducibility, or inter-run variability. In each run, three replicate standard curves were tested across the 384-well plate to assess repeatability, or intra-run variability. All reactions were performed in triplicates. Data analysis

Using the data generated, the following assay parameters were calculated: 1) inter-run assay coefficient of variation (CoV) for copy number and Ct value, 2) average intra-run assay CoV for copy number and Ct. value, 3) assay dynamic range, 4) average reaction efficiency, and 5) correlation coefficient (r 2 -value). The limit of detection was not defined for the pure plasmid standards selleck kinase inhibitor experiments due to variability in reagent contamination. At each plasmid standard concentration, the Ct standard deviation across all standard curves over three runs was divided by the mean Ct value across all standard curves over three runs to obtain the inter-run assay CoV. The CoV from each standard curve from each run (i.e., nine CoV were used in the calculation for each condition tested) were used to calculate the average and the standard deviation of the intra-run CoV. Linear regression of each standard curve across the full dynamic range was performed to obtain the slope and correlation coefficient values. The slope was used to calculate the reaction efficiency using Efficiency = 10(−1/slope)-1.

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