Design, development and synthesis of a novel labeled PNA monomer incorporated in DNA-hexamer to act as a hybridization probe by FRET

Abstract

A novel PNA monomer with adenine nucleobase and a modified backbone with charged <img src='/image/spc_char/nh.gif' border=0> and a methylene substituted for <img src='/image/spc_char/co.gif' border=0> in the linker arm has been synthesised. This modified PNA monomer is further linked with rhodamine via an amide bond, converted to phosphoramidite after linking with a 2-carbon linker. This unit is used for the end coupling at 5'-prime of a hexamer oligonucleotide 5'-TCC ATT-3' in solid phase. The complementary labeled heptamer has been prepared by coupling the phosphoramidite of fluorescein with 5'-AAT GGAT-3' as the end coupling step at 5'-prime on solid phase. The two complementary heptamer oligos are hybridized and the FRET signal recorded. The Tm of the modified oligomer has been found to be higher as compared to the normal one. Such a probe can detect single base mismatch by FRET