9 replies to this topic

[namekuji]

Is low melting point agarose gel supposed to be softer than regular agarose gel?  I made a 1.5% gel with low melting point agarose but it felt like 0.5% or something. It crumbled in my hands because I was not expecting such brittleness.  (Or is it our balance that's broken??)

[T C]

Hey

Its okay

It is like this

Best
TC

namekuji, on Mar 20 2009, 01:15 PM, said:

Is low melting point agarose gel supposed to be softer than regular agarose gel?  I made a 1.5% gel with low melting point agarose but it felt like 0.5% or something. It crumbled in my hands because I was not expecting such brittleness.  (Or is it our balance that's broken??)

[drevilette_R]

Really?? i personally dont think it s normal.. just yesterday i made a 2% gel and it felt fine.. i mean it s  not as hard as you normal agarose .. could it be that u used water instead of TAE?? (i know it s a stupid question ..but it happened to me once..so who knows..)
otherwise i can take a look at the brand that we re using if u re interested.
best regards

[T C]

Hey

It a different kind of agarose called LMP agarose and the gel is very fragile.  I generally use it for very small inserts like changing the MCS of a vector.  Also, the gel is transparent (partially).  

But I do agree with you that with companies, the quality of the agarose and thus the gel varies.  I have a bottle in the lab from SIGMA; when I use this agarose, the strength of a 1.5 % gel is same as that of a 1% gel with agarose from USB.



Best,
TC

drevilette_R, on Mar 20 2009, 04:23 PM, said:

Really?? i personally dont think it s normal.. just yesterday i made a 2% gel and it felt fine.. i mean it s  not as hard as you normal agarose .. could it be that u used water instead of TAE?? (i know it s a stupid question ..but it happened to me once..so who knows..)
otherwise i can take a look at the brand that we re using if u re interested.
best regards

[merlav]

In general all low melting agarose are fragile if compare with the regular one.  The LM should run  with a lower V than regular and is better to run at 4-10C.  They should be manage with care.  Try to add the etbr to the gel, because if stain after the running you could end with a puzzle.

[perneseblue]

If need to, I sometimes make a frame of normal agarose, which I then fill in with LM agarose. This frame of more solid agarose gives additional strength to the gel.

[C F]

perneseblue, on Mar 20 2009, 08:47 AM, said:

If need to, I sometimes make a frame of normal agarose, which I then fill in with LM agarose. This frame of more solid agarose gives additional strength to the gel.

My 1% gel fell apart by just taking out the comb!  Any suggestions on the preparation of the gel?
I made a standard 1% Agarose in 1x TAE, microwaved, poured, waited 20-30 mins and it came apart upon lifting up the comb.

[hobglobin]

C F, on Jun 1 2010, 09:27 PM, said:

perneseblue, on Mar 20 2009, 08:47 AM, said:

If need to, I sometimes make a frame of normal agarose, which I then fill in with LM agarose. This frame of more solid agarose gives additional strength to the gel.

My 1% gel fell apart by just taking out the comb!  Any suggestions on the preparation of the gel?
I made a standard 1% Agarose in 1x TAE, microwaved, poured, waited 20-30 mins and it came apart upon lifting up the comb.

Just be careful and lift the comb really vertical up...you can melt the gel and pour it again...this agarose is expensive...

[epibio]

It also helps to handle the gel (and run it, as someone suggested) in the cold room if you can.

[Avdin]

I've only ever used one type of LMP agrose so i don't know about other brands, but generally speaking I have found them to be much more brittle than regular gels. At voltages over 90 they begin to get even more fragile unless kept at a low temperature. What I have been able to do with great success is to use a little bit of regular agarose with the LMP agarose to stiffen it up. If I am making a 1.5% gel I will use 1.3% LMP and 0.2% regular agarose. I am actually currently trying to make a 1% gel in this same manner. Not sure how well it will work but I'm hopeful. The low amount of regual agarose does not seem to effect the gel extraction. The gel disolves nicely and I generally see what I estimate to be about 60-70% yeild.
Also, in order to speed up the solidification process and make sure it is as stiff as possible before removing the comb, I let it solidify in the fridge at 4 degrees for 15-30 minutes.