Daniel:Notebook/ComboLock/2017-5-16
Production Run (Started Friday 4/28; Libary Prep)[edit]
Sequencing Results[edit]
These results are from the sequencing run performed 5/8/2017.
Read Statistics[edit]
Sample | Condition | Reads Aligned | Reads Unaligned | Pct Aligned | Missing UMIs (%) | Poisson Lambda | Poissonian E[0] (%) | UMI Missing Enrichment |
1X | NEB Sample | 254728 | 21861 | 92.1% | 34.17 | 3.89 | 2.0 | 16.7 |
5AX | NEB Positive Control | 119220 | 10206 | 92.1% | 31.2 | 1.82 | 16.2 | 1.9 |
1Y | TF Sample | 197959 | 23195 | 89.5% | 42.53 | 3.02 | 4.9 | 8.7 |
5AY | TF Positive Control | 63074 | 6955 | 90.1% | 51.37 | 0.96 | 38.3 | 1.3 |
4AY | TF Dephosphorylated | 133660 | 15412 | 89.7% | 72.47 | 2.03 | 13.1 | 5.5 |
NA | Randomly Generated 250,000 UMIs | 2.26 | 3.81 | 2.21 | 1.0 |
UMI Counting[edit]
- Sample.neb-histogram.png
UMI count histogram from the NEB sample
- Sample.tf-histogram.png
UMI count histogram from the ThermoFisher sample
- Poscon.neb-histogram.png
UMI count histogram for positive control samples, from NEB
- Poscon.tf-histogram.png
UMI count histogram for positive control samples, from TF
- Dephos.tf-histogram.png
UMI count histogram for dephosphorylated samples (TF)
- RandomUMI.250K-histogram.png
UMI count histogram for randomly generated UMI list
Base Complexities[edit]
- Sample.tf-basecomplexity.png
Sample (TF) UMI base complexity
- Sample.neb-basecomplexity.png
Sample (NEB) UMI base complexity
- Poscon.tf-basecomplexity.png
Positive Control (TF) base complexity
- RandomUMIs-basecomplexity.png
Randomly generative UMI base complexity
Knee Plots (5/19/2017)[edit]
Added from analysis performed yesterday (5/18). The knee plots show the UMIs, ordered by number of reads (descending), vs. total fraction of reads. Only the random UMI samples go up to 1 on the y axis, but that's because the others don't have 100% alignment.
- Sample.neb-kneeplot.png
Sample (NEB)
- Sample.tf-kneeplot.png
Sample (TF)
- Poscon.neb-kneeplot.png
Positive control (NEB)
- Poscon.tf-kneeplot.png
Positive control (TF)
- Dephos.tf-kneeplot.png
Dephosphorylated sample (TF)
- RandomUMI.250K-kneeplot.png
Random UMI
Discussion[edit]
Sequencing results are good overall, at least good enough to move on to the BSA positive control and possibly cells for mRNA. Couple of notes:
- Production run positive control yielded correct sequences
- Samples had additional empty UMIs to theoretical Poissonian zero
- Dephosphorylated had most (72%) empty UMIs
- ThermoFisher had slightly better UMI complexity
- The knee plots show that the dephosphorylated sample had the highest AUC, which is actually bad for this type of analysis