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Help with positive control


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4 replies to this topic

#1 Sebastian Castillo

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Posted 24 October 2013 - 06:25 AM

Hi everyone. I am using methylamp (epigentek) for global DNA methylation quantification. This kit includes a DNA positive control for methylation, but I want to make my own. I ask for help in the last and they told me about pde9 gen who it's listed in de the paper of bock et al. Pubmed 16520826. I search in this papers and there is no sign of this gene.

So I still needs help, I need another positive control, this control can be a gene that it's full methylated, where I can find a list of possible candidates.
And what it's more useful, this or using a ssi methyltransferase kit to make my own methylated controls

#2 Sebastian Castillo

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Posted 24 October 2013 - 07:48 AM

In fact i search in methytilation database and that gene pde9 it's methylated in some samples and in others not.
Another possible gen?

#3 carthage

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Posted 29 October 2013 - 09:56 AM

Question -- can you describe why you intend to use a specific gene as the positive control for a global DNA methylation study?



#4 phage434

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Posted 29 October 2013 - 01:22 PM

You can make a fully methylated sample with PCR using 5me-dCTP instead of CTP. You can also methylate only the CGs of a DNA sample with M.SssI (NEB).



#5 et2b

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Posted 18 November 2013 - 01:52 AM

Indeed, full methylation of a gene often depends on the cell line or tissue you are looking in. Not a reliable 'control' . Go for enzymatic methylation for the 100% methylated and a whole genome amplified DNA for a 0%, you can than make a titration curve for your global meth analysis, much more insightful maybe....






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