Cleavage of Oligonucleotides from Solid Supports
Publication Date: 2004-Jul-14
The IP.com Prior Art Database
This disclosure describes processes for the cleavage of synthetic oligonucleotides from solid supports. These processes include an in-line method in which the cleavage reagents are pumped through the solid support bearing the oligonucleotide and contained in a column or similar vessel within in a reactor. The benefits of carrying out such in-line cleavage processes at elevated temperature are described.
Synthetic oligonucleotides, including chemically modified oligonucleotides containing one or more phosphorothiate residues, are often synthesized on a solid support while contained in a reactor or a fixed-bed column. Cleavage of oligonucleotides from solid supports (usually a polymer resin, or controlled pore glass) within a reasonable reaction time is a requirement for efficient synthesis. Usually, such cleavage processes are effected by treatment with a concentrated aqueous ammonia solution although alternative reagents such as methylamine or t-butylamine and/or cosolvents such as methanol and ethanol may also be employed. Such cleavage processes are normally performed by one of two modes: (a) as a batch-wise process following removal of the solid support bearing the oligonucleotide from a column or similar vessel within a reactor, or (b) as an in-line process in which the reagents effecting the cleavage are pumped through the solid support bearing the oligonucleotide while still contained in a column or similar vessel within in a reactor. For convenience and simplicity of operation, mode (b) can often be preferred. It can also be advantageous to conduct the in-line cleavage process at elevated temperature, provided that the equipment is properly designed to tolerate higher vapor pressures. Such benefit is described in the literature by Chow, in U.S. Patent no. 5496473:
“In order to speed up the removal of the DNA from the DNA synthesis column, means for controllably heating the columns are provided, e.g., a heating device that can be turned on during the clea...