added amplitude files from paper for example, updated script to make …

…figures and tables, added new information to readme

FAM_ABS_RT_coV.csv

@@ -0,0 +1,25 @@
+Well,ExptType,Experiment,Sample,TargetType,Target,Status,Concentration,Supermix,CopiesPer20uLWell,TotalConfMax,TotalConfMin,PoissonConfMax,PoissonConfMin,Positives,Negatives,Ch1+Ch2+,Ch1+Ch2-,Ch1-Ch2+,Ch1-Ch2-,Linkage,AcceptedDroplets,CNV,TotalCNVMax,TotalCNVMin,PoissonCNVMax,PoissonCNVMin,ReferenceCopies,UnknownCopies,Ratio,TotalRatioMax,TotalRatioMin,PoissonRatioMax,PoissonRatioMin,FractionalAbundance,TotalFractionalAbundanceMax,TotalFractionalAbundanceMin,PoissonFractionalAbundanceMax,PoissonFractionalAbundanceMin,ReferenceAssayNumber,TargetAssayNumber,Threshold,MeanAmplitudeofPositives,MeanAmplitudeofNegatives,MeanAmplitudeTotal,ExperimentComments,MergedWells,TotalConfMax68,TotalConfMin68,PoissonConfMax68,PoissonConfMin68,TotalCNVMax68,TotalCNVMin68,PoissonCNVMax68,PoissonCNVMin68,TotalRatioMax68,TotalRatioMin68,PoissonRatioMax68,PoissonRatioMin68,TotalFractionalAbundanceMax68,TotalFractionalAbundanceMin68,PoissonFractionalAbundanceMax68,PoissonFractionalAbundanceMin68
+"A01",Absolute Quantification,"ABS","Blank",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.34,0,0,10267,,,,,,10267,,,,,,,,,,,,,,,,,,1,1,3772,0,2912,2912,"",,,,0.16,0,,,,,,,,,,,,,
+"A02",Absolute Quantification,"ABS","10K_RT",Ch1Unknown,"FAM","Manual",2.1,One-Step RT-ddPCR Kit for Probes,42,,,3.1,1.3,20,11452,,,,,,11472,,,,,,,,,,,,,,,,,,1,1,3761,4983.9,3006.3,3009.7,"",,,,2.55,1.63,,,,,,,,,,,,,
+"A03",Absolute Quantification,"ABS","1K_noRT",Ch1Unknown,"FAM","Manual",0.1,One-Step RT-ddPCR Kit for Probes,2,,,0.48,0,1,11683,,,,,,11684,,,,,,,,,,,,,,,,,,1,1,3772,4787.5,2796.2,2796.3,"",,,,0.25,0.03,,,,,,,,,,,,,
+"B01",Absolute Quantification,"ABS","Blank",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.39,0,0,9106,,,,,,9106,,,,,,,,,,,,,,,,,,1,1,3772,0,2908.9,2908.9,"",,,,0.18,0,,,,,,,,,,,,,
+"B02",Absolute Quantification,"ABS","100K_RT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.34,0,0,10394,,,,,,10394,,,,,,,,,,,,,,,,,,1,1,3772,0,2978.6,2978.6,"",,,,0.15,0,,,,,,,,,,,,,
+"B03",Absolute Quantification,"ABS","1K_noRT",Ch1Unknown,"FAM","Manual",0.12,One-Step RT-ddPCR Kit for Probes,2.4,,,0.58,0.01,1,9755,,,,,,9756,,,,,,,,,,,,,,,,,,1,1,3794,4191.6,2769.6,2769.7,"",,,,0.3,0.03,,,,,,,,,,,,,
+"C01",Absolute Quantification,"ABS","Blank",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.37,0,0,9413,,,,,,9413,,,,,,,,,,,,,,,,,,1,1,3761,0,2907,2907,"",,,,0.17,0,,,,,,,,,,,,,
+"C02",Absolute Quantification,"ABS","100K_RT",Ch1Unknown,"FAM","Manual",0.11,One-Step RT-ddPCR Kit for Probes,2.2,,,0.51,0,1,11050,,,,,,11051,,,,,,,,,,,,,,,,,,1,1,3749,3903,2969.2,2969.3,"",,,,0.27,0.03,,,,,,,,,,,,,
+"C03",Absolute Quantification,"ABS","10K_noRT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.32,0,0,11001,,,,,,11001,,,,,,,,,,,,,,,,,,1,1,3727,0,2768.6,2768.6,"",,,,0.15,0,,,,,,,,,,,,,
+"D01",Absolute Quantification,"ABS","1K_RT",Ch1Unknown,"FAM","Manual",12.6,One-Step RT-ddPCR Kit for Probes,252,,,15,10.2,108,10042,,,,,,10150,,,,,,,,,,,,,,,,,,1,1,3761,4977.1,2994.5,3015.6,"",,,,13.8,11.4,,,,,,,,,,,,,
+"D02",Absolute Quantification,"ABS","100K_RT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.33,0,0,10720,,,,,,10720,,,,,,,,,,,,,,,,,,1,1,3749,0,2897,2897,"",,,,0.15,0,,,,,,,,,,,,,
+"D03",Absolute Quantification,"ABS","10K_noRT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.32,0,0,10930,,,,,,10930,,,,,,,,,,,,,,,,,,1,1,3761,0,2804.3,2804.3,"",,,,0.15,0,,,,,,,,,,,,,
+"E01",Absolute Quantification,"ABS","1K_RT",Ch1Unknown,"FAM","Manual",12.2,One-Step RT-ddPCR Kit for Probes,244,,,14.8,9.8,92,8860,,,,,,8952,,,,,,,,,,,,,,,,,,1,1,3761,4659.6,2791.8,2811,"",,,,13.5,10.9,,,,,,,,,,,,,
+"E02",Absolute Quantification,"ABS","1M_RT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.33,0,0,10733,,,,,,10733,,,,,,,,,,,,,,,,,,1,1,3761,0,3022.1,3022.1,"",,,,0.15,0,,,,,,,,,,,,,
+"E03",Absolute Quantification,"ABS","10K_noRT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.33,0,0,10773,,,,,,10773,,,,,,,,,,,,,,,,,,1,1,3716,0,2749.1,2749.1,"",,,,0.15,0,,,,,,,,,,,,,
+"F01",Absolute Quantification,"ABS","1K_RT",Ch1Unknown,"FAM","Manual",16.3,One-Step RT-ddPCR Kit for Probes,326,,,19.1,13.6,138,9875,,,,,,10013,,,,,,,,,,,,,,,,,,1,1,3727,4773.3,2887.3,2913.3,"",,,,17.7,14.9,,,,,,,,,,,,,
+"F02",Absolute Quantification,"ABS","1M_RT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.31,0,0,11240,,,,,,11240,,,,,,,,,,,,,,,,,,1,1,3761,0,2976.2,2976.2,"",,,,0.14,0,,,,,,,,,,,,,
+"F03",Absolute Quantification,"ABS","100K_noRT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.32,0,0,11013,,,,,,11013,,,,,,,,,,,,,,,,,,1,1,3738,0,2767.1,2767.1,"",,,,0.15,0,,,,,,,,,,,,,
+"G01",Absolute Quantification,"ABS","10K_RT",Ch1Unknown,"FAM","Manual",1.7,One-Step RT-ddPCR Kit for Probes,34,,,2.8,0.9,13,8999,,,,,,9012,,,,,,,,,,,,,,,,,,1,1,3761,4559.6,2797.5,2800,"",,,,2.21,1.27,,,,,,,,,,,,,
+"G02",Absolute Quantification,"ABS","1M_RT",Ch1Unknown,"FAM","Manual",0.12,One-Step RT-ddPCR Kit for Probes,2.4,,,0.56,0,1,9958,,,,,,9959,,,,,,,,,,,,,,,,,,1,1,3738,3860.7,2942.9,2943,"",,,,0.29,0.03,,,,,,,,,,,,,
+"G03",Absolute Quantification,"ABS","100K_noRT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.33,0,0,10562,,,,,,10562,,,,,,,,,,,,,,,,,,1,1,3749,0,2804.5,2804.5,"",,,,0.15,0,,,,,,,,,,,,,
+"H01",Absolute Quantification,"ABS","10K_RT",Ch1Unknown,"FAM","Manual",2,One-Step RT-ddPCR Kit for Probes,40,,,3.2,1.1,15,8801,,,,,,8816,,,,,,,,,,,,,,,,,,1,1,3783,4570.3,2818,2821,"",,,,2.6,1.5,,,,,,,,,,,,,
+"H02",Absolute Quantification,"ABS","1K_noRT",Ch1Unknown,"FAM","Manual",0.11,One-Step RT-ddPCR Kit for Probes,2.2,,,0.54,0,1,10456,,,,,,10457,,,,,,,,,,,,,,,,,,1,1,3738,6440.6,2791.1,2791.5,"",,,,0.28,0.03,,,,,,,,,,,,,
+"H03",Absolute Quantification,"ABS","100K_noRT",Ch1Unknown,"FAM","Manual",0,One-Step RT-ddPCR Kit for Probes,0,,,0.27,0,0,12880,,,,,,12880,,,,,,,,,,,,,,,,,,1,1,3772,0,2789.8,2789.8,"",,,,0.13,0,,,,,,,,,,,,,

README.md

@@ -1,4 +1,17 @@
 # Plotly and python implementation of define the rain
+http://definetherain.org.uk/
 
+# Instructions for use
 
-http://definetherain.org.uk/
+## Running ddPCR and export of files
+First run the ddPCR reactions using the BioRad QX200 machine and QuantaSoft™ software. Then export the Amplitude and Cluster Data to a .csv file. Place these files into a separate folder.
+
+## Kmeans clustering
+The software then creates a dataframe from all amplitude data. The user must then select a well that will be used to generate the clusters (a positive control that doesn't have 100% positive droplets)
+
+``` npposcontrol = df.loc[df['Sample'] == 'D02']['Ch1 Amplitude'].values ```
+
+After clustering the positive and negative cluster are used to generate positive and negative thresholds. The positive threshold is 3 standard deviations lower than the mean of the positive cluster. The negative threshold is 3 standard deviations higher than the mean of the negative cluster. Anything that is found between the two thresholds is call droplet rain.
+
+## Diagram and table generation
+Finally, a plotly tablet and diagram are generated from the amplitude data.