NSTI Nanotech 2009

Fluorescence Based Specific PNA-DNA Binding Characterization on nano-FET DNA Device Surface

S.K. Rastogi, N.N. Mishra, B. Finaloski, E. Cameron, W. Maki
Postdoc Researcher, US

Keywords: beta-Ala-T10 PNA, Synthesis, cDNA, liquid binding, surface binding, nano-FET device

Abstract:

A novel sequence of peptide nucleic acid (PNA; beta-Ala-(T10)-beta-Ala-NH2) containing ten thymine monomers and beta-alanine amino acid at both (C and N) terminal of main chain was synthesized manually using solid phase tBoc peptide chemistry. Synthesized PNA (beta-Ala-(T10)-beta-Ala-NH2) were analyzed and purified using RP-HPLC and characterized by MALDI-TOF mass spectrometry. The hybridization of beta-Ala-(T10)-beta-Ala-NH2 with its complementary single strand DNA (cDNA; A10) was established in liquid phase through non denaturing gel electrophoresis and UV-thermal denaturation curve at 1 M concentration. The Tm of hybrid was observed 40.05 C. The nano-FET devices were designed and fabricated in our group. The PNAs were covalently immobilized on device surface with alkyl-thiol self assembled monolayer. DAPI nucleic acid stain and fluorescine labeling methods were used to prove the binding of PNA with its cDNA on nano FET surface from 10 nM solution, using fluorescent microscopic imaging. beta-Ala-(T10)-beta-Ala-NH2 : A10 binding is very specific and didnt show any affinity with noncomplementry DNAs. The model reported here has a potential capability of applying in developing PNA based biosensors for specific nucleic acid detection at room temperature.
 
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