10/07/2014
Ran a gel of some of the samples from the qPCR plate to check for primer dimer products
Weird things about this gel:
- Bands in my no template control lanes
- Bands are fuzzy like it's not a single product
- Bands look too big
- Bands with the same primer set seem to be different sizes
Possibility 1: I made a mistake and added template to my no template controls
- Counter: The likely way I would do this would be to add a correctly diluted sample but the Cp of these wells is later than my lowest dilution on my curve
- Counter: Doesn't explain bands of different sizes for same primer set
Possibility 2: I'm only measuring primer dimer
- Counter: Why are my dilution curves so perfect? Seems the Cp is dependent on template concentration.
Sent PCR products for Luc14 gDNA Surveyor PCR products for each gRNA out for sequencing
| Sample
|
Primer
|
'
|
F/R
|
Distance from gRNA
|
| Luc14 gRNA 20 Srvyr PCR |
P165 |
gRNA 021 US P F |
F |
149
|
| Luc14 gRNA 21 Srvyr PCR |
P164 |
gRNA 020 US P R |
R |
166
|
| Luc14 gRNA 22 Srvyr PCR |
P165 |
gRNA 021 US P F |
F |
103
|
| Luc14 gRNA 27 Srvyr PCR |
P155 |
gRNA 026 ES P F |
F |
121
|
| Luc14 gRNA 29 Srvyr PCR |
P162 |
gRNA 030 ES P R |
R |
125
|
| Luc14 Srvyr PCR |
P165 |
gRNA 021 US P F |
F |
149
|
| Luc14 Srvyr PCR |
P164 |
gRNA 020 US P R |
R |
166
|
| Luc14 Srvyr PCR |
P155 |
gRNA 026 ES P F |
F |
121
|
| Luc14 Srvyr PCR |
P162 |
gRNA 030 ES P R |
R |
125
|
| 
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