Purification of oligonucleotides

GENERI BIOTECH provides 3 types of oligonucleotide purification:

  • Standard purification – final synthesis product is made free of chemical impurities only (excess of salts and protective groups). This type of purification is suitable for oligonucleotide lengths up to 40 – 50 bases.
  • OPC purification (Oligonucleotide Purification Cartridge) – final synthesis product is made free of chemical impurities (excess of salts and protective groups) and also of shorter incomplete chains. This type of purification is sufficient for most standard applications.
  • HPLC purification – oligonucleotide is very effectively separated from all impurities and improper-length sequences. HPLC purification provides highly pure oligonucleotide suitable even for the most demanding applications.We use as well a modified HPLC purification we have developed – HPLC2. It is a combination of ion exchange and reverse-phase HPLC with gradient elution, which potentiates the effectiveness of the purification and minimizes losses resulting from the repeated HPLC purification.
    HPLC2 increases quality of probes and modified oligonucleotides in such a way that when the same amount of probe is used in the reaction, it leads to fluorescence increase from 10 to 40% compared with probe purified by a standard HPLC.

Standard purification is not surcharged. Each oligo ordered with OPC or HPLC purification is surcharged with additional fee regardless delivery term, labelling or other modifications.

You can find out your prices of purified oligos using our oligo calculator (after you log-in to Customer Portal).

Comparison of probes purified by HPLC2 and our original HPLC

The graphs show the curves of qPCR probes designed for genotyping. Therefore, the qPCR curves do not have a textbook S shape. All experiments were performed on a real-time PCR cycler CFX96 / CFX96Touch (Bio-Rad) with corresponding GB probe and GB master mix.

Fig.1: Probe FAM-BHQ1
Green curve – corresponds to the recording of the probe after HPLC2
Blue line – probe purified by our original purification
RFU – relative fluorescence unit

 

 

 

Fig. 2: Probe HEX-BHQ1
Green curve – corresponds to the recording of the probe after HPLC2
Blue line – probe purified by our original purification
RFU – relative fluorescence unit

 

 

 

Guaranteed and usual yields of specified purification types (oligos 15-50 bases long)

SCALE 20 nmol Guaranteed yield (nmol) Usual yield (nmol)
standard purification 10 13-20
OPC purification 8 13-17
HPLC purification N/A N/A

 

SCALE 40 nmol Guaranteed yield (nmol) Usual yield (nmol)
standard purification 20 35-40
OPC purification 10 30-35
HPLC purification 5 10-35

 

SCALE 200 nmol Guaranteed yield (nmol) Usual yield (nmol)
standard purification 60 130-180
OPC purification 40 45-130
HPLC purification 20 25-100

 

SCALE 1 µmol Guaranteed yield (nmol) Usual yield (nmol)
standard purification 300 350-500
OPC purification 100 200-300
HPLC purification 50 100-250