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Method for phosphorothioate antisense DNA sequencing by capillary electrophoresis with UV detection
D Froim1, C E Hopkins, A Belenky
1Analytical Research, Hybridon, Inc., Cambridge, MA 02139, USA.
Nucleic Acids Research
|October 23, 1997
Summary
A new, cost-effective method for sequencing short single-stranded oligonucleotides using UV detection and capillary electrophoresis (CE) has been developed. This technique simplifies antisense DNA therapy analysis without expensive labeling.
Area of Science:
- Biochemistry
- Molecular Biology
- Analytical Chemistry
Background:
- Antisense DNA therapy requires efficient sequencing of short single-stranded oligonucleotides.
- Existing sequencing methods can be costly and complex, involving fluorescent or radioactive labeling.
Purpose of the Study:
- To develop a reliable, convenient, and cost-effective method for sequencing phosphorothioate antisense DNA.
- To adapt the chain termination sequencing method for UV detection coupled with capillary electrophoresis (CE).
Main Methods:
- A modified chain termination sequencing approach was employed.
- Capillary electrophoresis (CE) with UV detection was utilized for analysis.
- Sample processing was minimized, and no labeling was required.
Main Results:
- The method successfully generated sequencing products detectable by UV.
- Undesired components were discriminated by reducing their concentration below the UV detection limit.
- A 'clean' and well-defined sequence was obtained.
Conclusions:
- The developed CE-UV method offers a reduced-cost alternative for oligonucleotide sequencing.
- This technique simplifies analysis for antisense DNA therapy.
- The method is compatible with commercially available CE-UV equipment and automation.