Deb G.
>Hi,
>
>I would like to get some suggestions/feedback in regards to our
>purification method for
>"long" oligos (over 25bp). We have been having difficulty with low
>yield using a reverse
>phase cartridge (Waters Sep Pak C18). We synthesize the oligos leaving
>the last DMT "on" and our protocol is as follows:
>
>After cleavage and deprotection with concentrated NH4:
>
>1. Rinse column with 100% Methanol
>2. Rinse column with 0.1M Tea-Ac (Triethylamine / Acetic Acid pH 7.0)
>
>3. Load Oligo onto column with 0.1M Tea-Ac
>4. "Remove failures" with 11% Acetonitrile in 0.1M Tea-Ac
>5. Remove DMTs with 1.5% TFA
>6. Rinse with 0.1M Tea-Ac to neutralize TFA
>7. Rinse with dH20 to desalt
>8. Elute from column with ~ 1ml 40% Methanol, then lyophilized to
>dryness.
>
>Any suggestions or modifications to improve this protocol would be
>greatly appreciated.
>
>Thank you.
>Mark
>--
>Mark O. Lively, Ph.D.
>Professor of Biochemistry
>Wake Forest University School of Medicine
>Medical Center Blvd.
>Winston-Salem, North Carolina 27157
>Voice: 336-716-2969
>Fax: 336-716-7200
>email: mlively@wfubmc.edu
Deborah S. Grove, Ph.D.
Manager of the Nucleic Acid Facility
Biotechnology Institute
The Pennsylvania State University
210 Wartik Lab
University Park PA 16802
http://www.biotec.psu.edu/lsc/stf/naf/naf.html