Tag Archives: gel

Freeze and squeeze DNA extraction from gel

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Hi all,

Here’s a really easy DNA gel extraction:

1. Go about cutting out your bands of interest in the usual way.

2. Next cut the gel band(s) into tiny cubes.

3.  Place tiny cubes in an eppie.

4. Put eppie in -20oC for 15 to 30 min.  The idea is to make the band freeze into a mini ice cube.

5.  Knock out all of the gel band/cubes onto a 10 (long) x 5 (wide) cm piece of parafilm.

6. Fold the parafilm in half along the length while keeping your gel cubes along the fold in the parafilm.  You should have a piece of parafilm folded lengthwise with gel cubes sandwiched between two layers of plastic.

7.  Crush/squeeze the liquid out of the tiny cubes.  Liquid containing electrophoresis buffer and DNA will separate from the agarose.

8.  Collect DNA/electrophoresis buffer mixture with pipet and clean it by ethanol precipitation.

 

EDIT:  Borate in the electrophoresis buffer is a ligase inhibitor.  Make sure to thoroughly wash your DNA with 70% EtOH, at least 2 times.

Loading Dye for Agarose Gels (Dan E.)

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If you are new to the lab bench, and perhaps even if you’re not, you may not know how to make loading dye for agarose gels. Its the sort of thing that tends to get made rarely and then persist, so people often inherit a supply and have no idea how to make more when they run out. For this sort of thing the best reference is the relevant appendix at the back of the Sambrook et al. Molecular Cloning text books (near Megan’s desk) and, of course, there is Google. Or, you can just do what I do. My recipe is incredibly simple and would be close to the cheapest option too. Continue reading →

Gel doc Dropbox (Allan)

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Hi all,

I’m not sure how most of you were sharing your gels but I set up a dropbox folder within the Rieseberg gels folder.  Sending e-mails was tiresome.  If you’re interested in synchronizing your gels with your computer drag your folder(s) to the dropbox folder.  The dropbox account credentials (online and client) are as follows:

user=allandebono@allandebono.org

password=1d7hz@!

… Go through the process as usual to set up a shared folder.

Gel Slice – size selection or band isolation (Dan E.)

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Cutting a slice from an agarose gel in order to size select DNA or to isolate a PCR product etc. is standard practice in lab genetics. This post is about how I do it in our lab, (as of Feb 2012).

There are three features of my current method that may be unfamiliar to you even if you’ve done this sort of thing before:

  1. The Dark Reader transilluminator.
  2. SYBR-Gold.
  3. Sigma X-Tracta gel punches.

Continue reading →

Quantifying DNA (Dan E.)

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This post is about quantifying DNA samples in the Rieseberg lab, i.e. using the tools we have available to us in our lab in the Biodiversity building as of November 2011.

There are three ways to quantify the amount of DNA in an aqueous solution (e.g. DNA dissolved in water or TE):

  1. Nanodrop (spectrophotometry).
  2. Qubit (flourometry).
  3. Agarose gel with EtBr (also fluorometry -> UV lightbox + your eyes = fluorometer).
  4. (There is also the BioAnalyzer – I’m going to ignore that here and focus on methods to measure high mw genomic DNA).

These methods have different benefits and limitations but they are all valid and useful.
Continue reading →