User:Tara K. Luckau/Notebook/Team ConGen/2010/10/18

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 * style="background-color: #BCED91"|[[Image:owwnotebook_icon.png|128px]] Tara's Lab Notebook
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 * style="background-color: #9DB68C" align="center"|  |Main project page


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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!

Purpose

 * Rehydrate Scun2 primers for testing tomorrow
 * 1) Scun2-F
 * 2) Scun2-R
 * 3) Scun2-F-M13F(-21)
 * 4) 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

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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