Oligodeoxyribonucleotide Activation with 2,4-Phenylenediisothiocyanate and Their Covalent Grafting onto Amine-Functionalized Latex Microspheres

Abstract

The activation of various synthetic oligodeoxyribonucleotides (ODNs), bearing a hexamethylamine arm at their 5 0 -end, with the 2,4-phenylenediisothiocyanate (PDC) was studied. The activation reaction had to be performed within 40 min, because of competitive hydrolysis of the isothiocyanate functions. ODN modifications were then characterized by different analytical methods i.e. gel capillary electrophoresis, Matrix-Assisted Laser Desorption-Ionization-Time of Flight (MALDI-TOF) mass spectrometry. Moreover, activation of the amine groups of the nucleic bases was evidenced through the chemical analysis of the enzymatic-digested conjugate (PDC-ODN) by reverse phase HPLC. Preliminary results on the covalent grafting of activated ODN molecules onto the aminated latex particles showed that the amount of grafted ODNs was directly related to the number of potential grafting sites on the nucleic acid probe (primary amine group of hexamethyl arm and amine functions of the nucleic bases).

Functionalized Magnetic Emulsion for Genomic Applications

“…Then, various chemical grafting processes of oligonucleotides onto functionalized colloidal particles were investigated and reported [30,31].…”

Surface-functionalized latexes for biotechnological applications

“…Another approach consists in designing various types of reactive magnetic or nonmagnetic polymer particles bearing surfaceamino groups [68]. Experimental conditions were then optimized (mostly pH, ionic strength, presence of a nonionic surfactant) in order to ensure efficient chemical grafting of negatively charged oligonucleotides bearing an amino-terminal spacer introduced during the automated synthesis of the ODNs [69]. Thermally sensitive colloidal particles were found suitable for avoiding undesirable physical adsorption of the ODNs [70].…”