DNA Extraction from PAGE Gels

DNA Extraction from PAGE Gels
Applications for Oligonucleotide Purification

A well tried and established method to purify synthetic oligonucleotides, is the so called “soak and crush method”. Briefly the method, consist of:

1. After PAGE electrophoresis is completed, the oligonucleotide bands are visualized by the UV shadowing method

2. Using a scalpel, the oligonucleotide bands are carefully cut from the gel.

3. The bands are gently crushed and cut in to smaller fragments and placed into a test tube containing 5mls ( or more, but always enough to cover all the crushed gel)of 0.1 X TE(Tris EDTA)

4. The crushed gel is left for 3 hours at room temperature(or overnight if convenient; 37 degrees incubation, renders higher recovery and is optional.

5. A 1 cm slurry of DE-52 resin ( this is a positively charge resin, that will retain the polyanionic oligonucleotide) is prepared in a 10 cm plastic column with medium or coarse fritted filter

6. The liquid 0.1X TE,now containing the oligonucleotide is passed through the DE-52 bed

7. The column is washed 3x with 10mls each of dd water

8. Use a 1 molar solution of TEAB( triethylammonium bicarbonate buffer) to elute the oligo

9. Use 3x1 ml of the TEAB buffer

10. Using a speed vac, evaporate the TEAB buffer and measure your A260 absorbance; oligo is ready for your use

Bio-Synthesis.Inc, has been producing synthetic oligonucleotides for over 25 years; not only DNA, but RNA, other modified oligonucleotides, and provide cross-linking of various types of biomolecule using our optimized bioconjugation strategies, which have a number of applications on gene expression inhibition and related anti-sense studies. Also synthetic peptides and peptide antibodies for a number of biological relevant applications, in the areas of proteomics, epigenetics, immune regulation, post translational modifications, antisense, gene expression control, RNA interference and more. For more product information, visit: www.biosyn.com.