Re: [DIYbio] Re: PCR Question

On Fri, Jul 12, 2013 at 11:56 PM, Mega <masterstorm123@gmail.com> wrote:
> Hi,
>
> reviving this thread for one more question ;)
>
>
> I did a PCR of a 9-10 kbp plasmid and the product should have been some 6300
> bp long.
>
> Now on this gel you see two bands (+primers), one at 9kbp and one at 6 kbp.
> May this be template plasmid + product? Or is this template (circular
> plasmid) + template (supercoiled) ?

Which column are you questioning? You could have done a better
photoshop/GIMP/MsPaint job! Usually labels go above or below lanes,
not on the side with pointing lines.

See the first page here:
http://people.rit.edu/rhrsbi/GEPages/LabManualPDF5ed/15%20electrophoresis.pdf

basically a way to be sure is by cutting the plasmid initially.

But I think if you used the same amount of plasmid in the PCR reaction
as you did in the control lane (plasmid only) you could tell if the
PCR worked simply by the difference in band brightness!

I'm actually not sure what any of your lanes are, which is the ladder,
which ladder is it? Did you use a ladder (prepared standard size
fragments)?

>
> Best,
> Andreas
>
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--
-Nathan

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