Standard curves for environmental samples

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Standard curves for environmental samples

Postby et124 » Mon Oct 25, 2010 8:59 pm

What is the best way to create a standard curve for the analysis of a gene in environmental samples? I am interested in running qPCR that will quantify a gene in a taxonomic group (taxa-specific primers, not species specific).

How similar does the sequence I use in my standards have to be to the sequences in my environmental samples? If the sequences of my target gene in my environmental samples vary from the sequence in my standards will I still get a valid number? Is it best to create the standards from the sequence from a culture organism? Will I be able to do this using degenerate primers?

Thank you!
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Joined: Mon Oct 25, 2010 8:47 pm

Re: Standard curves for environmental samples

Postby Mikael Kubista » Fri Oct 29, 2010 6:15 am

If the purpose of the standard curve is to provide estimates of target concentrations in field samples then it is critical the standards are as similar to the field samples as ever possible. This holds for the target sequence s well as for the sample matrix. Any systematic difference will lead to a bias in the concentration estimates.

Good luck
Mikael Kubista
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Joined: Tue Jul 01, 2008 12:28 pm

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