| Summary: | 碩士 === 中國文化大學 === 應用化學研究所 === 95 === ABSTRACT
The exploitation of hairpin-type triplex formation has been a versatile and rational approach to sequence-specific DNA recognition. In this thesis, two deoxyoligonucleotide probes 5'-d-(AG)4T4(CT)4 and 5'-d-(TC)4T4(CT)4 were designed to recognize the target strands 5'-d-(TC)4 and 5'-d-(AG)4, respectively. A variety of ionic compounds, including Na2SO4, NH4Cl, NaCl, (NH4)2SO4, CH3COONa, HCOONa and MgSO4 in a concentration range of 0.2-2.0 M, were tested in a phosphate buffer of pH 6.0 for the ability to modulate the structural stability of the triplexes 5'-d-(AG)4T4(CT)4+5'-d-(TC)4 and 5'-d-(TC)4T4(CT)4+5'-d-(AG)4, as well as their respective core duplexes. The structural stability were identified by UV thermal melting study and the Tm values were determined from the melting profiles.
Experimental results revealed that the Tm values of the triplexes 5'-d-(AG)4T4(CT)4+5'-d-(TC)4 and 5'-d-(TC)4T4(CT)4+ 5'-d-(AG)4 were elevated as the concentrations of ionic compounds Na2SO4, NH4Cl, NaCl or (NH4)2SO4 were increased from 0.2 M to 2.0 M.
In the sulfate solutions (0.2-0.4 M), the cationic effect on thermal stability of the above two triplexes showed an order: Mg2+ > NH4+ > Na+. However, it gave varied trends in the concentrations of 1.0-2.0 M. In the chloride solutions (0.4-2.0 M), the cationic effect was also shown NH4+ > Na+.
On the other hand, in sodium ion solutions (0.2-2.0 M), the anionic effect on stabilization of 5'-d-(AG)4T4(CT)4+5'-d-(TC)4 or 5'-d-(TC)4T4(CT)4+ 5'-d-(AG)4 followed an order: SO42- > Cl- > HCOO- > CH3COO-. In the same range of 0.2-2.0 M with the cation NH4+, the enhancement by SO42- was also more effective than that of Cl-.
It was observed that the triplex 5'-d-(TC)4T4(CT)4+ 5'-d-(AG)4 gave single melting temperature under conditions studied. Thus, the dissociation of the Watson-Crick domain and the Hoogsteen domain had almost the same Tm value.
Thermodynamic results by van't Hoff calculation appeared that free energy of the triplex decreased as the melting temperature increased.
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