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How many fragments in plasmid DNA prep?


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#1 science noob

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Posted 29 June 2012 - 03:37 PM

How many bands would you get if you run an agarose gel with a prep product from a transformation experiment?

I've used the ethanol precipitation method to derive my plasmid.

Heard that DNA comes in three forms - supercoiled, circular and linear. Can someone enlighten me on this? Cheers!

#2 bob1

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Posted 29 June 2012 - 05:41 PM

Ideally you would get all three, in practice, you will probably also get some digested/fragmented DNA (small mw smear, could also be RNA) and some genomic DNA which will be large, near the top of the gel/in the wells.

The three forms are the native state (supercoiled - tightly wound and may run faster or slower than circular, but usually faster), circular (single strand break in the plasmid which allows supercoil to unwind, or enzymatically unwound) and linear (double strand break, runs at actual plasmid size).

For transfections of eukaryotic cells you want supercoiled DNA preferably, apart from in a few specialized applications.




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