DNA "refusing" to be run in agarose gel

Ole Skovgaard olesk at RUC.DK
Fri Aug 25 06:52:19 EST 1995


In article <ketilt.4.000D5FD6 at dmf.unit.no> ketilt at dmf.unit.no (Ketil Thorstensen) writes:
>From: ketilt at dmf.unit.no (Ketil Thorstensen)
>Subject: DNA "refusing" to be run in agarose gel
>Date: Fri, 25 Aug 1995 13:22:22

>Honored contributors to the molbio.methds-reagnts!

>I have a peculiar problem with persuading some of my DNA samples to run in 
>agarose gels.

>When loading DNA samples on agarose gels I have on several occasions observed 
>a strange (to me) phenomenon: after application of a sample into a well a 
>streak of the sample may follow the pipette tip upon withdrawal. When the 
>sample in this way comes in contact with the buffer surface, the sample very 
>rapidly spreads out to cover the entire surface. During this process most of 
>the sample in the well is "sucked" out.
>There is no recognizable pattern as to what sample this may happen with. It 
>is, however, frustrating when precious samples are lost (the last one was a 
>1.9 kb DNA probe)  :í-(
>Could anyone enlighten me as to what I am observing, and how to prevent 
>this from happening again?

>Thank you very much,

>Ketil Thorstensen, Ph.D.
>Dept. Clinical Chemistry
>University Hospital
>Trondheim, Norway
> 
It is a well known phenomenon. It is due to the presence of either detergent 
or organic solvent i.e. ethanol/isopropanol from the last precipition before. 
Do your precipitation but remove ALL - repeat ALL - liquid before 
continuation. Do it with add of a pipette, centrifuge and remove the remaining 
liquid. Do not do it by inverting the tube or other silly things that 
biochemists have taught us (accept my apology for being root). Do not dry the 
pellet. Do not wash the pellet. That makes it go away - just remove all 
liquid!! and redisolve in whatever buffer. If there has been phenol in the 
tube change it immediately. Quite an amount of phenol may hide in the tube 
wall.
Add enough loading buffer for the 
gel to increase density of your sample above that of the gel buffer. Good luck,
ole


******************************************
****   Ole Skovgaard                  ****
****   Dept. Life. Sc. & Chemistry    ****
****   Roskilde University, DK        ****
******************************************



More information about the Methods mailing list