User:Tara K. Luckau/Notebook/Team ConGen/2010/10/18: Difference between revisions
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* H6, H7 evaporated, but I think it's workable | * H6, H7 evaporated, but I think it's workable | ||
* Plan to do first optimization PCR tomorrow! | * 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 | |||
* (C<sub>1</sub>) (V<sub>1</sub>) = (C<sub>2</sub>) (V<sub>2</sub>) | |||
*: (2.5mM) (5000µL) = (100mM) (V<sub>2</sub>) | |||
*: V<sub>2</sub> = 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 H20 from room 325 | |||
* Aliquots: 1000µL into each of 5 snap-caps; 4 stored in freezer, 1 stored in fridge for immediate use | |||
Revision as of 17:17, 28 October 2010
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PCR Thermalcycler Testing2 Mats w/ TapeDescription
Results
Primer Rehydration, Scun2Purpose
Calculations
dNTP MixPurpose
Calculations
Wet Work
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