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The red Low Read II samples were renormalized via pooling and the new MC samples were added to that same pool at 1 ul per sample. This pool was then adjusted to 1nM. The pool of the repeated RMJan22 samples was also adjusted to 1 nM. 50 ul of Low Read was added to 100 ul RMJan22. However, when the LowRead pool was qPCRed, one replicate was much higher than the other 2. The 1:2 ratio might be off because of this. |
RNaseP Check of qPCR machine recalibration:
Recalibration looks good. A snapshot is below:
Sample Name | Detector | Task | Ct | StdDev Ct | Qty | Mean Qty | StdDev Qty |
|---|---|---|---|---|---|---|---|
5K | RNase P | Unknown | 27.80 | 0.052 | 5170.54 | 4935.10 | 168.205 |
5K | RNase P | Unknown | 27.80 | 0.052 | 5164.36 | 4935.10 | 168.205 |
10K | RNase P | Unknown | 26.77 | 0.058 | 10193.14 | 9923.36 | 380.987 |
10K | RNase P | Unknown | 26.82 | 0.058 | 9863.28 | 9923.36 | 380.987 |
NTC | RNase P | NTC | Undetermined | ||||
NTC | RNase P | NTC | Undetermined | ||||
NTC | RNase P | NTC | Undetermined | ||||
NTC | RNase P | NTC | Undetermined | ||||
Standard1 | RNase P | Standard | 29.94 | 0.039 | 1250.00 | ||
Standard1 | RNase P | Standard | 29.92 | 0.039 | 1250.00 | ||
Standard1 | RNase P | Standard | 30.00 | 0.039 | 1250.00 | ||
Standard1 | RNase P | Standard | 30.00 | 0.039 | 1250.00 | ||
Standard2 | RNase P | Standard | 28.92 | 0.059 | 2500.00 | ||
Standard2 | RNase P | Standard | 28.87 | 0.059 | 2500.00 | ||
Standard2 | RNase P | Standard | 28.82 | 0.059 | 2500.00 | ||
Standard2 | RNase P | Standard | 28.78 | 0.059 | 2500.00 | ||
Standard3 | RNase P | Standard | 27.81 | 0.034 | 5000.00 | ||
Standard3 | RNase P | Standard | 27.82 | 0.034 | 5000.00 | ||
Standard3 | RNase P | Standard | 27.86 | 0.034 | 5000.00 | ||
Standard3 | RNase P | Standard | 27.88 | 0.034 | 5000.00 | ||
Standard4 | RNase P | Standard | 26.91 | 0.013 | 10000.00 | ||
Standard4 | RNase P | Standard | 26.93 | 0.013 | 10000.00 | ||
Standard4 | RNase P | Standard | 26.90 | 0.013 | 10000.00 | ||
Standard4 | RNase P | Standard | 26.89 | 0.013 | 10000.00 | ||
Standard5 | RNase P | Standard | 25.76 | 0.062 | 20000.00 | ||
Standard5 | RNase P | Standard | 25.69 | 0.062 | 20000.00 | ||
Standard5 | RNase P | Standard | 25.67 | 0.062 | 20000.00 | ||
Standard5 | RNase P | Standard | 25.61 | 0.062 | 20000.00 |
Assign reads and otus to samples:
...
Returns: 15371232
Divided by 8: 1921404 assigned to samples.
Assigned/Total (*100) = percent assigned: ~73%
...
Divided by 8: 1412494
LRII + locad2: 1921404
Even if all the unassigned reads are from locad2, this does not fix the expected ration of 2lo:1LR.
[508910+(2630678-1921404)] = 1218184 total possible locad2 reads
...