04/22/2015
Objective: Ran pcr with template cDNA created from 4 rna sea pen rna samples on 04/15/2015 with Sam at the Roberts lab
used following temp. settings for the thermalcycler


Conventional PCR (2x Apex Red) [Cost per rxn ~ $0.52]
Single reaction (25uL) set up is listed below. Be sure to make a master mix volume that will accommodate all of your samples, two water (no template controls; NTC) samples, plus an extra 10% to accommodate pipetting errors. Distribute appropriate amount of master mix (volume of master mix + template = 25uL) to PCR tubes or PCR plate. Make sure all tubes/caps are tightly closed. Put in thermalcycler.
Reaction_Components
Volume
Final Concentration
2x Apex Red
12.5
1x
Forward Primer (10uM)
0.5
0.2uM
Reverse Primer (10uM)
0.5
0.2uM
Template
Up to 5uL

H2O (PCR grade)
variable
Use to bring reaction volume up to 25uL
Typical cycling paramaters (ask for help on using the thermal cycler):

95C - 10mins
40 cyles of:
95C - 15s
55C - 15s
72C - 1 mins (dependent on amplicon size; ~1000kb/min)

modified quantity measurements

Results: Sucessfully completed PCR will run out on gel to veirfy replication of cDNA.