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PCR Probe Primer Design

PCR Probe Primer Design - PCR - Polymerase Chain Reaction Forum

PCR Probe Primer Design - PCR - Polymerase Chain Reaction Forum. Discuss and ask questions about PCR troubleshooting, PCR protocols and methods, PCR products, and PCR theory.


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Old 04-14-2009, 09:13 PM
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Question PCR Probe Primer Design



I'm trying to get primers/probes for MSRA-PBP ..I've used a couple of these free sites. Why are the primers all different? I'm trying to get primers/probes for a mulitplex PCR for MRSA. I want to cover the Scc-ORFx region, but I don't knop how to do it. I'm sure I'm missing something because I am so new at this. THX

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Old 04-18-2009, 04:22 PM
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Default Re: PCR Probe Primer Design

The results of your primer searches are different using different soft because they use different algorithms. There are better and worse programs to do so but always be careful to check your primers additionally for specificity (use BLAST) and quality (check the possibility of primer-dimers formation - especially for multiplex).
When you do multiplex you'll need to check whether the mixture of the primers amplify only your target sequences or maybe some unspecific as well. This is much easier (a lot easier ) if the genome of the organisms you're working on has already been sequenced and is present in databases [this is likely the case with S.aureus]. Well, it sometimes happens that when you try your primers, despite their carefull selection, you'll have to redesign them. Fortunately the primers are not very expensive.
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