Many different applications demand different purity or scale to work well when it comes to custom DNA oligo synthesis. Here are some guidelines for purity and synthesis scale selection for different applications.
Understanding Why Oligos Sometimes Require Purification


Purification Options Offered

Purification Method Description Benefit
(25 nm: 10-100 bp; 50 nm: 5-100 bp)
Oligos are processed through normal phase chromatography column which removes salts but not failure sequences. A salt-free DNA solution, ready-to-use; suitable for many PCR and sequencing applications without further purification.
(50 nm – 1 µm, 7-55 bp)
Based on reverse phase chromatography; removes failure sequences from the completed synthesis. Provides full-length sequences needed in some applications
(50 nm+, 10-55 bp)
Reverse Phase High Performance Liquid Chromatography (HPLC) removes failure sequences or unincorporated label the same way as cartridge purification. Guarantees highly purified primer required in some applications (>=85% full length).
(200 nm+, 7-100 bp)
Method used to differentiate full-length product from failure sequences based on size and conformation. Provides the highest percentage of full-length oligos (>=85%) required for certain demanding applications such as mutagenesis or adapter production.

Application-Purity Guide

Application Suggested Purity
AFLP™ AnalysisDesalted oligos have been used successfully for Amplification Restriction Fragment Polymorphisms.
First-Strand cDNA Synthesis for Generation of LibrariesGenerally oligos for first strand cDNA synthesis for library construction has some sequence at the end which codes for 5´ restriction endonuclease cloning sites. Therefore, it is best to use full-length, Cartridge, HPLC, or PAGE-purified oligos.
Fluorescent SequencingAll four purity grades have worked successfully for Invitrogen scientists.
Gel Shift AssaysCartridge, HPLC, and PAGE-Purified oligos are recommended for gel shift assays, so as to have a homogeneous population of DNA fragments.
GENETRAPPER® ScreeningPAGE-purified oligos are recommended. Primers should be phenol extracted and ethanol precipitated prior to use in the tailing reaction in GeneTrapper® System. If Desalted Purity oligos are purchased they can be PAGE-Purified using the PAGE purification protocol.
Isothermal SequencingDesalted oligos are sufficient for this application, along with Cartridge, HPLC, and PAGE-purified.
MicroarraysStandard desalted oligos are sufficient for printing onto arrays.
Production of Cloning AdaptersFull-length oligos work best for efficient cloning. Utilize cartridge, HPLC, or PAGE-Purified oligos for full length.
Site-Directed MutagenesisFull-length (e.g., Cartridge, HPLC, and PAGE-purified) oligos as a rule tend to give the highest percentage of mutagenized clones (especially if the intended mutation is close to the 5´ end of the oligo). Desired mutations have been obtained using Desalted oligos. However, some wild-type parental vector clones tend to carry over.