Electrochemical Characterization and Hybridization Efficiency of Aminated DNA Probes Immobilized on Screen Printed Carbon Electrodes
| Main Author: | Wijayanti, SudarmaDita |
|---|---|
| Format: | Thesis NonPeerReviewed |
| Terbitan: |
, 2011
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| Subjects: | |
| Online Access: |
http://repository.ub.ac.id/157705/ |
Daftar Isi:
- The research has focused on the how to attach DNA strands to carbon screen printed electrodes via an amino group, so that they stand up with a good orientation. This is to ensure the full length of the probe is available for hybridization. Electrodeposition at -1.0 V and covalent attachment of the aminated DNA probes by 1-ethyl-3-(3 dimethylaminopropyl)-carbodiimide (EDC) and N - Hydroxysuccinimide (NHS) . The DNA coverages and hybridization efficiencies were determined using both chronocoulometry and UV absorption. The use of propylamine as a spacer for the DNA strands, the effect of the propylamine : DNA ratio and the effect of the probe length were also examined. The best results were gained by immobilising pDNA-15 spacing with propylamine spacer at a 0.5 mole fraction ratio. Maximum ssDNA coverages determined by chronocoulometry were 1.37 x 10 13 molecules cm -2 obtained for electrodeposition and 4.97 x 10 12 molecules cm -2 for covalent attachment. The hybridization efficiencies ranged from 39.9 % to 80.3 %. These coverages were lower than those obtained by UV absorption. Measurement by UV-vis absorption does not involve the charge compensation assumptions required by the chronocoulometric method. The affinity constant (Ka) for duplex formation for the probes immobilized by covalent attachment ( 1.52 x 10 8 M -1 ) was higher than for the electrodeposition method ( 1.51 x 10 7 M -1 ) . Presumably, covalent attachment gave the ssDNA better orientation on the electrode in terms of allowing hybridization.