formaldehyde gel for RNA

gordon allison goa at aber.ac.uk
Thu Jun 20 06:31:10 EST 1996


In article <4q51m2$5cm at nuscc.nus.sg>, zoowyw at leonis.nus.sg says...
>
>Dear netter,
>
>I'd tried to run my RNA sample on formaldehyde agarose as protocol on
>Molecular cloning by Sambrook several times, but I get smear bands for
>samples and marker and failed to get two distinct band for 28S and 18S as
>expected. I'd eliminated the possibility of RNase contamination. Is these
>caused by quality of formaldehyde or pH?  Can I have references for these
>problem? 
>
>Thanks in advance.  
>Jason 


If you follow the method and it doesn't work then either you've done it 
wrong or else your RNA is crap. Points to note are:

1	Make sure that your formaldehyde is ok and doesn't contain too much 
polymerised lumps.
2	Denature the RNA in the formamide/ formaldehye loading buffer for 
15 mins at 65 C and then snap on ice.
3	I tend to run low formaldehyde gels now at about 0.6 M.
4	 You can put etidium in the loading buffer of stain the gel later 
in 0.5 ug/ ml ethidium in water or acetate. I generally transfer and stain 
the blot with methylene blue.

Good luck, Gordon.




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