Introduction

PNA is a kind of DNA analog that can recognize and combine DNA or RNA sequences to form a stable double-helix structure.

Fmoc is used as the protecting group of α-amino of amino acid. Its advantage is that it is stable under acidic conditions which are not affected by trifluoroacetic acid (TFA) and other reagents, and can be deprotected by mild alkali treatment. At the same time, the Fmoc method has fewer side reactions, and high yield, and the Fmoc group itself has characteristic UV absorption, which is easy to monitor and control the reaction.

Synthesis Steps

The amino backbone of Fmoc PNA monomers is protected by Fmoc, which enables PNA to bind to peptides and markers such as biotin or fluorescent dyes. Each synthesis cycle takes about 30 minutes, mainly including the following steps:

1. Install the column. Unlike the conventional peptide synthesis column that connects the synthesized C-terminal amino acid to the solid phase support, PNA synthesis column only contains polystyrene solid phase support.
2. Washing. Washing PNA synthetic column with dimethylformamide (DMF)
3. De close. Removal of Fmoc protective group with DMF
4. Washing. Remove excess reagent by washing with DMF.
5. Activation and coupling. In the presence of a mixture of diisopropylethylamine and hexafluorophosphate, the newly synthesized PNA monomer is activated and coupled to the solid phase resin.
6. Washing. Wash with DMF to remove excess reagent.
7. Cap. The unreacted gene is capped by acetic anhydride dissolved in DMF.
8. Washing. Wash with DMF to remove excess reagent.
9. The PNA is incubated with trifluoric acid and m-cresol mixture for cutting and final deprotection.

Application

Fmoc compound is selected to protect PNA monomer. After one monomer reacts at each point, its synthesis efficiency is tested by bromophenol blue color reaction. The efficiency of the synthesized PNA oligomer can reach 95%.