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Agarose gel cutting


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8 replies to this topic

#1 shimshady

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Posted 18 March 2009 - 07:30 AM

I am terrible at cutting my dna from an agarose gel. I dice it up pretty much and still have large pieces of agarose without dna. Do you have any advice for cutting up the gel with minimal agarose for purification? Thanks

#2 perneseblue

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Posted 18 March 2009 - 08:11 AM

I am terrible at cutting my dna from an agarose gel. I dice it up pretty much and still have large pieces of agarose without dna. Do you have any advice for cutting up the gel with minimal agarose for purification? Thanks


What do you use to cut your gel?

I use a fresh scalpel blade. But I only press down into the gel. I don't drag the blade in a cutting motion to avoid damaging the glass surface of the transluminator.
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#3 labrat612

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Posted 18 March 2009 - 10:39 AM

I too use a fresh scalpel but I also put a piece of plexiglass in between my gel and the transilluminator to avoid breaking the glass.

#4 hobglobin

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Posted 18 March 2009 - 12:01 PM

I reduce the UV intensity as far as possible and switch the lamp off immediately when not needed (we have an additional lamp to have enough light in the dark room).
Practice helps a lot, I was quite bad too, therefore I cut several times DNA ladders not needed anymore.
Here some just mark with a first "preliminary" cut the area they need and then cut on the unilluminated surface besides the glass.

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#5 bob1

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Posted 19 March 2009 - 12:12 AM

I do the preliminary cut as well. I have also noticed that the majority of the DNA is at the bottom of the wells, unless you mix just after loading each well. This means that you can cut the top of the slice off to minimise the amount of agarose.

#6 AquaPlasmid

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Posted 14 May 2009 - 10:00 PM

The most critical thing about DNA fragment purification is UV damage and crosslinking, if you use b-agarase digestion there is not a big deal with a little extra gel. If I am gel purifying a fragment for cloning, particularly for library cloning, I load a wide prep gel well and before I expose the gel to UV, I cut 3/4 off the gel lane and never expose it to UV. I only view the other 1/4 of the lane and cut a line on the DNA band with a raze blade. Then I turn off the UV and align the two gel pieces and cut out the band under lab light so my DNA fragment used for ligation will never see the UV light. If you have problem with ligation and have trouble getting your clone, try this trick.

#7 jiajia1987

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Posted 21 May 2009 - 12:21 AM

to make my life easier, i make use of an transilluminator without UV that still allows me to view my gel and cut them without the lights being on.

#8 hanming86

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Posted 21 May 2009 - 03:35 AM

to make my life easier, i make use of an transilluminator without UV that still allows me to view my gel and cut them without the lights being on.



Are u saying that u don't use UV to view Etbr stained DNA??
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#9 bob1

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Posted 21 May 2009 - 04:21 PM

You can get blue light transilluminators for DNA visualisation with fluorescent dyes such as sybr safe.




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