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Short Oligonucleotide DNA Microarrays :

Short Oligonucleotide arrays are created using in-situ photolithographic synthesis

25-mer oligos are chemically synthesized onto glass slides using photolithographic methods pioneered originally by Affymetrix.  Digitally controlled micro-mirrors are now used in the photolithographic printing technique.

Glass slides are coated with synthetic linkers which contain photo-labile chemical groups.  Masks are used to direct light to pre-determined positions on the slide and this removes exposed groups. The unprotected groups can then be coupled with bi-functional deoxy-nucleotide triphosphates dNTPs.

New masks are used in order to direct coupling at other sites until the desired sequences and lengths of oligos are synthesized.

An Affymetrix chip can have over 500,000 25-mer oligo probes arrayed on a 1.25x1.25 cm slide!

Anywhere from 10 to 20 different oligo probes are synthesized for each gene to tile from the 3' end of the mRNA.  There are oligos that are synthesized as perfect matches (designated PM), and mis-match pairs (MM).

Each MM pair has a one-base pair mismatch in the centre of the oligo, which allows a control for background noise and allows normalization of cross-hybridization.

 

Problems of Short Oligonucleotide DNA Microarrays:

- Short oligos have cross-hybridization problems

- Must therefore use many oligo controls for each gene target