Hello everyone,
Im performing an absolute quantification assay using real time PCR, and for this i am constructing a standard curve with linearized plasmid DNA. Does any one know how to prepare log 10 fold dilutions of the template DNA.?
pls help
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#2
ivanbio
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Posted 21 May 2010 - 08:58 PM
This may not be what you are looking for, but after reading your question I could not help but think this is the simple answer: just take 10 ul of your undiluted template and and mix it with 90 ul of water. That is a 10-fold dilution. Then take 10 ul of this 10-fold dilution and mix it with 90 ul of water to make a 100-fold dilution. Repeat this until you have 5 dilutions, which you can use as templates for your standard curve.
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Ivan
Carlsbad, CA
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vladooo
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Posted 21 May 2010 - 11:26 PM
ivanbio, on May 22 2010, 06:58 AM, said:
This may not be what you are looking for, but after reading your question I could not help but think this is the simple answer: just take 10 ul of your undiluted template and and mix it with 90 ul of water. That is a 10-fold dilution. Then take 10 ul of this 10-fold dilution and mix it with 90 ul of water to make a 100-fold dilution. Repeat this until you have 5 dilutions, which you can use as templates for your standard curve.
Cheers
Cheers
Exactly. And vortext vigorously between steps.
BTW: start with 10e8 copies of your plasmid
