Abstract
Diblock copolymers composed of allele-specific oligodeoxyribonucleotide (ODN) and poly(ethylene glycol) (PEG) are used as an affinity probe of free-solution capillary electrophoresis to quantitatively detect single-base substitutions in genetic samples. During electrophoresis, the probe binds strongly to a wild-type single-stranded DNA analyte (WT) through hybridization, while it binds weakly to its single-base-mutated DNA analyte (MT) due to a mismatch. Complex formation with the probe augments the hydrodynamic friction of either analyte, thereby retarding its migration. The difference in affinity strength leads to separation of the WT, MT, and contaminants, including the PCR primers used for sample preparation. The optimal sequence of the probe's ODN segment is rationally determined in such a way that the binding constant between the ODN segment and MT at the capillary temperature is on the order of 10(6) M(-1). The validity of this guideline is verified using various chemically synthesized DNA analytes, as well as those derived from a bacterial genome. The peak area ratio of MT agrees well with its feed ratio, suggesting the prospective use of the present method in SNP allele frequency estimation.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.