Fluorescence Based PNA-DNA Binding Characterization on Nano-Field Effect Transistor (FET) DNA Device Surface

摘要

A sequence of peptide nucleic acid (PNA) containing ten thymine monomers and β-alanine amino acid at both (C and N) terminal of main chain (βAla-(T)_(10)-βAla-NH_2) was synthesized manually using solid phase tBoc peptide chemistry. Synthesized PNAs were analyzed and purified using RP-HPLC and characterized by MALDI-TOF mass spectrometry. The hybridization of βAla-(T)_(10)-βAla-NH_2 with its single strand complementary DNA (sscDNA; A_(10)) was established in liquid phase through non denaturing gel electrophoresis and UV-thermal denaturation curve at 1 uM concentration. The T_m of hybrid was observed 40.05 ℃. Further, the nano-FET devices were designed and fabricated by our group for developing DNA biosensor. The PNAs were covalently immobilized on device surface with alkyl-thiol self assembled monolayer. DAPI nucleic acid stain and fluoresceine labeling methods were used to prove the binding of PNA with A_(10) on nano FET surface from 10 nM solution, using fluorescent microscopic imaging. The model reported here has a potential capability of applying in developing PNA based biosensors for specific nucleic acid detection at room temperature.