How to perform gradient PCR?

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w19910411
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How to perform gradient PCR?

Post by w19910411 » Wed Mar 11, 2015 8:47 am

How should one perform gradient PCR, and how much time will it take?

Javedz2001
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Post by Javedz2001 » Sun Oct 11, 2015 4:54 pm

The time will remain the same. Set up PCR as follows:

FAST Cycling Conditions Temp (°C) Time (sec) Initial denaturation/Hot Start 95 30 Steps 1–3 are repeated through 40 cycles
Step 1: 95 for 10 seconds
Step 2: 54–70 (gradient) for 15 seconds
Step 3: 72 15-60 seconds (depends on the length of your amplicon)

Good Luck: Javed Siddiqi

Fornita
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Re:

Post by Fornita » Mon May 23, 2016 4:19 am

Javedz2001 wrote:The time will remain the same. Set up PCR as follows:
Step 2: 54–70 (gradient) for 15 seconds
Good Luck: Javed Siddiqi


For step 2, you could change the temp. gradient according to your sequences properties. lower of higher.
Andrea Fortina
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CreativeBiomart, Recombinant protein expert

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