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 Tara's Lab Notebook
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PCR Thermalcycler Testing
2 Mats w/ Tape
Description
- 10µL in each well of full-skirt PCR plate, PCR seal, 2 silicone mats, lab tape on opposite sides
- Data: 20102018_TKL_CyclerTest2MatTape.tad
Results
- H6, H7 evaporated, but I think it's workable
- Plan to do first optimization PCR tomorrow!
Primer Rehydration, Scun2
Purpose
- Rehydrate Scun2 primers for testing tomorrow
- Scun2-F
- Scun2-R
- Scun2-F-M13F(-21)
- Scun2-R-pigtail
Calculations
- Scun2-F
- 28.3nmol + 141.5 Low TE = 200µM STK
- Scun2-R
- 29.2nmol + 146µL Low TE = 200µM STK
- Scun2-F-M13F(-21)
- 26.6nmol + 133µL Low TE = 200µM STK
- Scun2-R-pigtail
- 30.5nmol + 152.5µL Low TE = 200µM STK
- stored overnight in fridge to rehydrate completely before making dilution to 20µM
dNTP Mix
Purpose
- Mix dNTPs and aliquot for PCR
Calculations
- Have: Invitrogen 100mM dNTP Set (catalog# 10297-018) (250µL of 25µmol in H2O @ 100mM each)
- Want: 10mM dNTP mix (2.5mM each), 5 aliquots of 1000µL
- (C1) (V1) = (C2) (V2)
- (2.5mM) (5000µL) = (100mM) (V2)
- V2 = 125µL of each dNTP
Wet Work
- 125µL dATP + 125µL dTTP + 125µL dCTP + 125µL dGTP + 4500µL H2O = 5000µL dNTP Mix @ 10mM (2.5mM each dNTP)
- Used NanoPure H2O from room 325
- Aliquots: 1000µL into each of 5 snap-caps; 4 stored in freezer, 1 stored in fridge for immediate use
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Tara's Lab Notebook
