problems with background doing in vitro transcription/translation

Arthur Wohlwill U55850 at uic.edu
Sat Jun 22 23:49:15 EST 1996


In article <v01530500adf9d4425d4c@[142.20.8.21]> jlight at SICKKIDS.ON.CA writes:
>From: jlight at SICKKIDS.ON.CA
>Subject: problems with background doing in vitro transcription/translation
>Date: 28 Jun 1996 10:35:09 -0700

>I'm using the TNT coupled reticulocyte lysate system from Promega for in
>vitro transcription/translation, and I'm getting a lot of background signal
>at the top and bottom of my autorads.  I can understand that the signal at
>the bottom would be unincorporated 35S-methionine which I can run off the
>gel.  However, why the signal at the top?  The reason that this is a
>problem is that my product has a high MW and is obscured by the background
>at the top.  Also, I encounter background smearing in many of my sample
>lanes, although the positive control lane looks nice.  Any advice on how to
>get rid of background with this assay would be appreciated.  Thanks in
>advance. Jeff.


Sometimes there are complexes with charged tRNA's----Adding RNAse may help.
Arthur Wohlwill adwohlwi at UIC.EDU






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