Parallel preparation of high fidelity probes in an array format

A method of fabricating a plurality of oligonucleotides having free 3′-hydroxyl groups from a high density oligonucleotide array, said method comprising the steps of

• a) providing a solid substrate comprising a plurality of ribonucleotides attached thereto, one said ribonucleotide shown below
• wherein PG1 is protecting group 1, PG2 is protecting group 2, B is a naturally or non-naturally occurring base, and said ribonucleotide is attached to said substrate through the 5′-hydroxyl group;
• b) selectively removing PG1 in pre-selected areas to provide a plurality of free 3′-hydroxyl groups on said ribonucloetide;
• c) reacting said free 3′-hydroxyl groups with a 2′-deoxyribonucleotide having the structure
• wherein PG3 is protecting group 3 and RG is a reactive group to couple said 2′-deoxyribonucleotide to said ribonucleotide to provide the structure
• d) selectively removing PG3 from the 5′-hydroxyl of said 2′-deoxyribonucleotide in pre-selected areas to provide a plurality of free 5′-hydroxyl groups;
• e) reacting said free 5′-hydroxyl groups with an additional 2′-deoxyribonucleotide having the structure
• to yield a product of the structure
• f) repeating steps d and e one or more times to provide said oligonucleotides attached to said solid substrate;
• g) removing PG2 from one or more of said ribonucleotides to provide a free 2′-hydroxyl group on each of said one or more ribonucleotides;
• and
• h) transesterifying each of said one or more ribonucleotides to yield said solid substrate having a cyclic ester attached thereto and free oligonucleotides, each oligonucleotide having a 3′-hydroxyl group and having the structure