I don't know what you call "low yield" and which synthesis scale you use
but I think your protocol is right. What I have experienced with Sep Pack
cartridge (but it was for quite lipohilic methyl phosphonate oligos) was
always a certain lost of material (something like 30-50% of irreversible
interactions ) from 7 OD to 3 OD (18 mers) after dealting on Sep pack. I
heard about it was quite common even for normal phosphodiester to loose
some material on this kind of phase.
Sincerely,
Alain
>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
**************************
Alain LAURENT Ph. D
Oligonucleotide Chemistry
ESGS Groupe CYBERGENE
11 rue Claude Bernard
35400 SAINT MALO
FRANCE
Tel: +33 (0)2 99 21 90 40
Fax: +33 (0)2 99 21 90 41
e-mail: a.laurent@eurosequence.com
http://www.eurosequence.com
***************************