Overview
To establish some quality control standards for the RNA-seq and qPCR experiments, I performed qPCR on all dilutions of all templates using mCherry & GAPD as targets.
Templates to be used are:
- ✓ U2OS (1:10, 1:100, 1:1000)
- ✓ KAH126-U2OS (1:10, 1:100, 1:1000)
- SK-N-SH (1:10, 1:100, 1:1000)
Also will be trying out a duplex reaction with GAPD & mCherry primers in the same sample, using a template that shows strong mCherry signal.
qPCR plate layout for U2OS & KAH126-U2OS templates
qPCR results for U2OS & KAH126-U2OS templates
Experiment: DualColorProbe_UPL_DNyer042215 Active filters: FAM (465-510), VIC / HEX / Yellow555 (533-580)
Chart
|
Pairing
|
Sample Name
|
Targets
|
References
|
Mean Cp
|
Mean Cp
|
Target/Ref
|
Status
|
TRUE |
A1/A4 |
U2OS_E003 1:10 |
mCherry |
GAPD |
20.0372991 |
20.11290542 |
1.054 |
|
TRUE |
A7/A10 |
U2OS_E003 1:100 |
mCherry |
GAPD |
22.86651753 |
23.18434575 |
1.246 |
|
TRUE |
B1/B4 |
U2OS_E003 1:1000 |
mCherry |
GAPD |
26.63819532 |
27.28223381 |
1.563 |
|
TRUE |
B7/B10 |
U2OS_C003 1:10 |
mCherry |
GAPD |
29.20480103 |
19.51357142 |
1.21E-03 |
|
TRUE |
C1/C4 |
U2OS_C003 1:100 |
mCherry |
GAPD |
28.94313572 |
22.16621867 |
9.12E-03 |
|
TRUE |
C7/C10 |
U2OS_C003 1:1000 |
mCherry |
GAPD |
29.09594237 |
23.68882936 |
2.36E-02 |
|
TRUE |
D1/D4 |
U2OSKAH126_E001_A 1:10 |
mCherry |
GAPD |
17.62458299 |
18.98810117 |
2.573 |
|
TRUE |
D7/D10 |
U2OSKAH126_E001_A 1:100 |
mCherry |
GAPD |
21.64213569 |
22.82095575 |
2.264 |
|
TRUE |
E1/E4 |
U2OSKAH126_E001_A 1:1000 |
mCherry |
GAPD |
24.29214814 |
25.91215397 |
3.074 |
|
TRUE |
E7/E10 |
U2OSKAH126_E001_B 1:10 |
mCherry |
GAPD |
18.51985613 |
19.7082754 |
2.279 |
|
TRUE |
F1/F4 |
U2OSKAH126_E001_B 1:100 |
mCherry |
GAPD |
20.61966384 |
21.8860216 |
2.406 |
|
TRUE |
F7/F10 |
U2OSKAH126_E001_B 1:1000 |
mCherry |
GAPD |
24.82426956 |
26.35374426 |
2.887 |
|
TRUE |
G1/G4 |
U2OSKAH126_C001 1:10 |
mCherry |
GAPD |
23.51659697 |
19.4546205 |
5.99E-02 |
|
TRUE |
G7/G10 |
U2OSKAH126_C001 1:100 |
mCherry |
GAPD |
26.19546455 |
22.47707261 |
7.60E-02 |
|
TRUE |
H1/H4 |
U2OSKAH126_C001 1:1000 |
mCherry |
GAPD |
28.40332046 |
26.65931589 |
0.2985 |
|
|