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sequencing

Manuel G. CLAROS claros at biologie.ens.fr
Wed Dec 8 08:28:19 EST 1993


In Article <2e3dqc$42d at apakabar.cc.columbia.edu>,
ger4 at merhaba.cc.columbia.edu (Gabrielle E. Rieckhof) wrote:
>Dear bionetters,
>
>This is not a yeast question, per se, but I am sequencing a yeast gene. Anyway,
>when I sequence, I routinely load one set of sample, and load a second set 
>after the xylene cyanol marker of the first set has run out of the gel. When I
>am ready to load the second set, however, the top of the gel (the part that the
>comb touches to form the wells) has become distorted. I do not know if this is
>due to the heat or the electrical current, but it interferes with loading of 
>the second set of samples.
>
>I've found that running the gel on a lower wattage helps, though not 
>completely. I am using a wedge spacer on a 6% gel, and running at 50 or less 
>watts (about 1200V and 40mA). 
>
>Thanks in advance for any and all help.
>
>KC.
>
>P.S. Another problem I have is that the gel also developes air bubbles several 
>centimeters from the top after extended runs. Any remedies?
>
I had had this kind of problem. But it seemed localised in the particular gel
because others do not had it. You can forgive me if I say to you that I 
run my gels at 63W (1900-2300V, near 50 degrees) and use not to have
problems. However, try to change acrylamide or Tris.

Manuel
-------------------------------------
!  Manuel G. CLAROS                 !
!  Ecole Normale Superieure - CNRS  !
!  Lab. Genetique Moleculaire       !
!  46 rue d'Ulm                     !
!  75005 Paris (France)             !
!  E-mail: claros at biologie.ens.fr   !
!  Fax:    331-44 32 39 41          !
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