RNA gels - good protocol required (Sep/26/2006 )

Hi,
I've noticed that my RNA gels are ugly. good clear results, they're just ugly. instead of having nice straight bands (aka, what you should see), the RNA seems to clump together and i get squishy, blobs. it's not a smear, or anything like that... the results are very obvious, not a problem with my RNA or anything. they resemble a DNA gel which is horribly overloaded.... sorta. the blob, but smaller, and more precise.

the protocol i use is :

dH20 - 55.5mL
agarose - 0.75g
10X MOPS - 7.5mL
37% formaldehyde - 12mL

i melt the water and the agarose in the microwave, cool it down, then add the mops, then add the formaldehyde. the tank where it is run is with 1x mops. i run it at 90V for 30-40 minutes.

the gel feels squishy to touch. what am i doing wrong? anyone had this problem? i would like to see a clean band, not a clean squishy blob.

V

This is our recipe for RNA gels.

40ml
= 34 ml of H2O DEPC
= 400 mg agarose
= 4ml of 10x MOPS
= 2ml Formaldehyde 37%



10 x MOPS - 500ml

0.2 M MOPS 21gm MOPS
50 mM Na Acetate 8.35 ml of 3 M solution
10 mM EDTA 10 ml of 0.5 M
pH = 7.0

Loading Buffer for RNA
To be prepared new everytime.

400ul
= 250ul of Formamide
= 83ul of Formaldehyde 37%
= 50 ul of 10x MOPS
= 8 ul of 0.5 % Bromphenol
= 7ul of Eth. Bromide

Yaarrr Vetticus

Check out my protocol on the BioTechniques forum

http://molecularbiology.forums.biotechniqu...pic.php?t=10217