We had a similar case recently. 5'-15 RNA bases - 18 DNA bases-3'. Our
advantage is that the trityl analysis is shown on the computer screen as a bar
graph (Expedite 8909 Workstation). What happened was the DNA part went OK
(synthesis goes 3' to 5'), coupling efficiency was close to 100%, but with each
of the first 5 RNA bases the trityls dropped to unacceptably low level (I can
E-mail you the picture). Surprisingly, the machine did not stop (despite the
fact that the trityl failure alarm is working for regular DNA and should
normally halt/abort the synthesis). When we ran a DENATURING PAGE gel, sure it
was a 19-22 bases long product instead of the expected 33, nothing below and
nothing above. We e-mailed the pictures (trityls and PAGE) to the amidite
supplier and asked for an exchange of the faulty RNA amidites for other stuff
we buy from them (we got that), and then we dropped them as a supplier of
amidites. Needless to say, this happened in spite of all precautions we took
for work with RNA.
My advice - change your supplier for amidites.
Regards,
victor
www.alphadna.com
> -----Original Message-----
> From: Bao-Shiang Lee <boblee@uic.edu>
> To: Recipients of ABRF List <abrf@aecom.yu.edu>
> Date: 4 mars, 1999 02:17
> Subject: Chimer
>
> >
> >Hi everyone,
> >
> >I am making a Chimer (RNA & DNA; 5'29DNA+11RNA+6DNA+10RNA+15DNA 3') for a
> >user. However, the product is only 34mer. Any suggestions are appriciated.
> >
> >Best regards,
> >
> >Bob Lee