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Re Lost T's

Graham Graham
Thu Feb 13 09:28:16 EST 1997


Hi Everyone
							A month or so ago I posted a question about my lack of T peaks.  
I've had several helpfull personal replys for which I am very 
grateful.  It did occur to me that it may be better to send them to 
the list so that everyone could see them though.

		Well at last it seems I've found the problem.  It all began with the 
377's major snag, red rain.
	
	We were suffering so we reduced the temperature to 48 dec C.  We did 
this manually for the prerun and the run.  So, after seeking advice, I 
set up a new run module in which the temperature was changed to 48 deg 
C.

	This was a duplicate of the original, and only the temperature was 
changed.  Well it seems that the machine just did not like it.  I've 
even done the ultimate test,  run samples on a regular module with the 
temperature changed manually,  then run those same samples on my new 
module.  

	Wouldn't you know it I got a problem,  but it was not only the T's 
that were missing,  some traces lacked A's and T's,  while others 
lacked C's and yet others had not G's.  This is all on the same gel!

	Anyway I'm satisfied that this was the problem,  and changing the 
temperature manually gives no problems at all.

				As for the red rain, I've found the advice on this list exteamly 
helpfull.  I now put a wet towel over the end of the gel about 5 to 10 
minutes after it's poured.  I do wait until the acryalmide has  
definately set before I do it. 

	I've also tried (once) to pour the gel using ABI's gel pouring 
gizzmo, which I figure should keep the bottom from drying out.  This 
worked very well.  Now I must see if I can run the gels at 51 deg C!

	The red rain 'phenomenon' appears at the top of the gel even when I 
run at  48,  so I'm still not realy sure what causes it.

												Anyway thanks for all your help, and I hope this will be 
of use to someone.

						Graham Watson
							ICRF
							ADL



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