length of primer in PCR

sarahrahim · Post by **sarahrahim** » Fri Oct 17, 2008 10:48 pm

length of primer in PCR

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Post by sarahrahim » Fri Oct 17, 2008 10:48 pm

explain why the length of PCR primer is always approximately 20 - 30 oligomers? Is it possible to use a primers pair with about 10 oligomers? Explain the answer

stopherlogic · Post by **stopherlogic** » Fri Oct 17, 2008 10:54 pm

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Post by stopherlogic » Fri Oct 17, 2008 10:54 pm

Well why do you think? Do you have any ideas?

sarahrahim · Post by **sarahrahim** » Fri Oct 17, 2008 11:04 pm

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Post by sarahrahim » Fri Oct 17, 2008 11:04 pm

is it because the fact that dna polymerase can grab around 19 oligomer primer?

does it related to annealing temperature that related to the length of primer?

that might cause us to have non-specific amplification if we use too long a primer or a short one?

stopherlogic · Post by **stopherlogic** » Fri Oct 17, 2008 11:13 pm

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Post by stopherlogic » Fri Oct 17, 2008 11:13 pm

You thought of more than I did, the only reason I could think of was too short a primer and you would end up amplifying too many sequences in the genome. This is because the primer sequence would be non specific to the site that you want to amplify.

Don't quote me on it though.

sarahrahim · Post by **sarahrahim** » Fri Oct 17, 2008 11:19 pm

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Post by sarahrahim » Fri Oct 17, 2008 11:19 pm

no, i am new and really need help. this is my test question that my lecturer asked us to find out. he said this is too simple. i don't know how to explain if anybody have some more specific answer will be very appreciated.

mith · Post by **mith** » Fri Oct 17, 2008 11:50 pm

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Post by mith » Fri Oct 17, 2008 11:50 pm

The annealing temperature is related to the specificity of the sequence because of energy considerations. Note that at higher temperatures it's easier to make a non-favorable interaction i.e. wrong base pairing. However, if you have a long enough sequence, it will become unfavorable enough to make it only bind to specific sequences. You also want to consider whether you're running PCR on an unknown sequence i.e. leave room for mutations.

Secondly, you have to consider whether you're going to bind to a different part of the genome with the same sequence(dependent on length). This is a probability argument.

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samfisher · Post by **samfisher** » Fri Oct 24, 2008 7:10 am

Re: length of primer in PCR

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Post by samfisher » Fri Oct 24, 2008 7:10 am

Hi all

It is important issue that the length of primer in PCR.

But please if you explain the success of RAPD-PCR which use oligo decamere nucleotides as primer and there were no misspriming or false priming.

Q2: If there is a software to design RAPD primer?

sam fisher