Video Protocols > How to perform colony PCR
How to perform colony PCR
By Alan M. 2105 Views
Intermediate
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Summary
Here's an easy way to rapidly screen ligation reactions for positive clones - this tip will change your molecular biology life. Allart Stoop showed me, now I'm showing you.
Products Used in This Protocol
100x20 cell culture dish
Supplier: BD Biosciences
Catalog Number: 353003
200uL TipOne yellow tip wafers
Supplier: USA Scientific
Catalog Number: 1111-0706
Ampicillin, sodium salt
Supplier: SigmaAldrich
Catalog Number: A9518-5G
FailSafe PCR 2X PreMix, D
Supplier: Epicentre
Catalog Number: FSP995D
P20 Pipetman
Supplier: Gilson
Catalog Number: F123600
P200 Pipetman
Supplier: Gilson
Catalog Number: F123601
PCR 8-well Tube strips and caps
Supplier: VWR
Catalog Number: 20170-004
Round-bottom culture tubes, 15mL
Supplier: BD Biosciences
Catalog Number: 352059
SealRite eppi tubes, 1.5mL
Supplier: USA Scientific
Catalog Number: 1615-5500
Taq DNA Polymerase
Supplier: New England Biolabs
Catalog Number: M0273S
UltraPure Agarose
Supplier: Invitrogen
Catalog Number: 15510-027
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Comments (5)
"Hi, Alan. How much primer do you use for each reaction? Isn´t necessary boil the colony or just rubbing the tip in tube is a safely way?"
- Mariz, FC, Jul 12 @ 9:51 am
"Hi, Alan. How much primer do you use for each reaction? Isn´t necessary boil the colony or just rubbing the tip in tube is a safely way?"
- Mariz, FC, Jul 12 @ 9:39 am
"Excellent point Jane- no particular reason. I think you could go either way (colony or mix first). Saving tips and thumb pain is always a plus!"
- Alan M., Feb 3 @ 11:46 am
"Tomorrow I'll do my first colony PCR... I hope it will be so easy as you show us.
Nice video!"
- Guille, Jan 20 @ 10:25 pm
"I find it interesting that you transferred the colony into the tube prior to adding the master mix into each tube. Is there a reason why you do it that particular way? When you do it that way, you have to change the tip twice for each colony (when transferring the sample and adding the mix). However, by adding the mix first, you can use the same tip for each tube."
- Jane D, Dec 3 @ 1:21 pm