Synthesis of Photolabile 5′-O-Phosphoramidites for the Photolithographic Production of Microarrays of Inversely Oriented Oligonucleotides

Abstract

The photolabile 3′‐O‐{[2‐(2‐nitrophenyl)propoxy]carbonyl}‐protected 5′‐phosphoramidites (16 – 18) were synthesized (see Scheme) for an alternative mode of light‐directed production of oligonucleotide arrays. Because of the characteristics of these monomeric building blocks, photolithographic in situ DNA synthesis occurred in 5′→3′ direction, in agreement with the orientation of enzymatic synthesis. Synthesis yields were as good as those of conventional reactions. The resulting oligonucleotides are attached to the surface via their 5′‐termini, while the 3′‐hydroxy groups are available as substrates for enzymatic reactions such as primer extension upon hybridization of a DNA template (see Fig. 2). The production of such oligonucleotide chips adds new procedural avenues to the growing number of applications of DNA microarrays.

Personalised proteome analysis by means of protein microarrays made from individual patient samples

“…Basically, any number of proteins could be studied, although our work dealt with still relatively few molecules to date. In combination with the ability to synthesise oligonucleotides in situ on array surfaces in 5′ to 3′ direction in a directed and quantitative manner17, even each individual array could be different to the next for a truly targeted and individualised analysis. Nothing of this kind could be achieved at a reasonable cost and effort by individually purified proteins.…”

Organometallic DNA–B12 Conjugates as Potential Oligonucleotide Vectors: Synthesis and Structural and Binding Studies with Human Cobalamin‐Transport Proteins

“…Oligodeoxynucleotide synthesis: General : Pharmacia LKB Gene Assembler Special; the fast (Tac)deprotection approach was used . HPLC : Dionex Ultimate 3000 pump, with variable wavelength detector, column and autosampler column compartments.…”

DNA Microarray Preparation by Light‐Controlled In Situ Synthesis

“…Nearly all of these photolabile protecting groups are typically employed at the 5′ position, thereby enabling the preparation of DNA microarrays that expose a free 5′ end while leaving the 3′ terminus attached to the solid support. More recently, DNA microarrays that expose a free 3′ hydroxyl group have gathered special interest, opening the way for a new generation of applications that make use of enzymatic on‐chip reactions (Beier et al, ). This unit focuses on experimental conditions that have been tested for the in situ synthesis of oligonucleotide microarrays using NPPOC‐protected phosphoramidites (Fig.…”