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Blank Gel

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Blank Gel

Postby Tokophyra125 » Tue Jan 29, 2008 2:38 pm

In my Cell and Molec. lab last week we ran 0.8% gels with samples of RNA from fruit flies. A total of five lanes were loaded with a ratio of 2 /10 for the loading dye to samples. The gel was run at 75 V. for 45 mins. We were told to leave the lab and that the TA's would handle it from there and take pictures of the gels so we could do our write-ups. Upon checking my e-mail early this morning I discovered that absolutely nothing showed up on my group's gel. Not even the ladder was present, which I thought was odd. I think all the samples made it into the wells and didn't float out. So I am wondering what exactly happened?
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Postby mith » Tue Jan 29, 2008 3:06 pm

wrong polarity?
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Postby canalon » Wed Jan 30, 2008 2:10 am

A few possibilities:

-RNAse in the gel: all degraded.
-wrong polarity: everything went out through the top in a few minutes.
-The TA screwed up: you are screwed up...
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Postby MichaelXY » Thu Feb 07, 2008 9:25 pm

Wouldn't the wrong polarity just cause the direction in the gel to be reversed.
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Postby blcr11 » Thu Feb 07, 2008 9:33 pm

Sure would--and all your nucleic acid sample would migrate up out of the well into the upper chamber and you would see *nada* on the gel.
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Re: Blank Gel

Postby MichaelXY » Thu Feb 07, 2008 9:40 pm

Now don't I feel foolish. :oops:
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Re: Blank Gel

Postby blcr11 » Thu Feb 07, 2008 9:43 pm

Should add, I was assuming the gel was an open-well, top-loaded gel. If it was a horizontal gel with the wells to one end, if the polarity was wrong you had maybe 15 minutes to reverse the polarity back to the correct one before the samples ran off the end of the gel. It's generally a good idea to watch the dye fronts for the first several minutes of a run to make sure eveything is running correctly.
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Postby blcr11 » Thu Feb 07, 2008 9:45 pm

Nothing to feel foolish about. If you do these types of gels, I can pretty much guarantee you will reverse the polarity at least once in you lifetime.
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Postby MichaelXY » Fri Feb 08, 2008 9:07 pm

Thanks Blcr, I have only had 1 gel lab so far, and in our exp, the wells were in the middle of the gel. We added dyes to the wells and noted some dyes migrated towards pos end while others to the neg term. So i guess I better wait until the end of the semester before I spout off on this sort of thing. I thought it was a cool lab though. The prof said that phoresis is what crime labs use. cool stuff I thought. Thanks for your considerate response.
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Postby n0vad3m0n » Fri Feb 15, 2008 11:58 pm

We ran a gel in my AP bio class a few weeks ago. In our group's gel, we were missing several bands for one of our DNAs and through some intense stain/de-staining we could barely make some out.

Another possibility would be that you might have punctuated the gel and the samples ran over the place.
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