title: Automated purification and quantification of oligonucleotides.
abstract: We have developed automated methods for the trityl-on purification and quantification of synthetic oligonucleotides. Oligonucleotide purification is by solid-phase extraction cartridges using Amberchrom CG-50 resin on an XYZ-axis robotic system. Quantification is by OD260nm using an online UV-visible spectrophotometer with sipper. The purification of 20 oligonucleotides requires 5 min of user set-up time, plus 20 min per sample of robot time. For a 15-25-mer at the 40 nmol scale of synthesis, the method gives a yield of 2.8 ODs from a load of 10.1 OD, i.e., a 28% average yield. Oligonucleotides purified by this method have proven to be successful for primers for automated DNA sequencing.
authors.affiliation: Biomolecular Resource Center, University of California, San Francisco 94143, USA. kathyi@cgl.ucsf.edu
authors.forename: K M
authors.initials: KM
authors.lastname: Ivanetich
authors.forename: R C
authors.initials: RC
authors.lastname: Reid
authors.forename: R
authors.initials: R
authors.lastname: Ellison
authors.forename: K
authors.initials: K
authors.lastname: Perry
authors.forename: R
authors.initials: R
authors.lastname: Taylor
authors.forename: M
authors.initials: M
authors.lastname: Reschenberg
authors.forename: A
authors.initials: A
authors.lastname: Mainieri
authors.forename: D
authors.initials: D
authors.lastname: Zhu
authors.forename: J
authors.initials: J
authors.lastname: Argo
authors.forename: D
authors.initials: D
authors.lastname: Cass
authors.forename: C
authors.initials: C
authors.lastname: Strickland
mesh_terms: D001322:Autoanalysis; D001483:Base Sequence; D002845:Chromatography / Q000191:economics / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D003365:Costs and Cost Analysis; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000032:analysis* / Q000138:chemical synthesis / Q000302:isolation & purification; D010957:Plasmids; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013056:Spectrophotometry, Ultraviolet; D013698:Templates, Genetic; D013997:Time Factors
chemical_list: D009841:Oligonucleotides