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McClean: Sequencing Colony PCR Product: Difference between revisions
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McClean: Sequencing Colony PCR Product (view source)
Revision as of 12:27, 25 March 2013
, 25 March 2013no edit summary
(New page: <!-- COPY EVERYHING BELOW HERE TO START YOUR OWN PROTOCOL! --> ==Overview== This protocol is for sequencing the product of a yeast colony PCR. ==Materials== * Colony PCR product * DNA ...) |
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==Materials== | ==Materials== | ||
* Colony PCR product | * Colony PCR product | ||
* DNA Clean & Concentrator (Zymo Research) | * DNA Clean & Concentrator Kit (Zymo Research) | ||
* Sequencing primers | * Sequencing primers | ||
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# Elute your DNA (Step #5) in 10μL of H<sub>2</sub>O (NOT Elution Buffer). This is the eppendorf tube you will send for sequencing, so make sure it is clearly labeled. | # Elute your DNA (Step #5) in 10μL of H<sub>2</sub>O (NOT Elution Buffer). This is the eppendorf tube you will send for sequencing, so make sure it is clearly labeled. | ||
# Use 2μL of this DNA product to check concentration on the nanodrop. | # Use 2μL of this DNA product to check concentration on the nanodrop. | ||
# To the 8μL of remaining DNA, add 4μL of primer. | # To the 8μL of remaining DNA, add 4μL of primer at 8pmol. (Our lab oligos are at 10μM, so to get the desired 8pmol of oligo, dilute our oligos 1:5 in sterile water for a final of 2pmol/ul and add 4μL to the 8μL of DNA to be sequenced). | ||
