投影片 1

•Oligonucleotides

–usually at least 20–25 bases in length, optimal with 45~60 bps long

–10–20 different oligonucleotides for each gene

•Oligonucleotides for each gene selected by computer program to be the following:

–unique in genome (4 (20 to 25) =2(40 to 50) >> 3*109 = 230), not likely to appear twice

–non-overlapping (if the sequence length is too short then specificity is low, whereas if the length is too long, self-hybridization could happen)

•Composition based on design rules

–empirically derived rules

(ratio of G-C pairs vs. A-T pairs which

could affect the melting temperature

of the seq., ie. Tm = 64.9+0.41*(GC%)

-675/L)

Oligonucleotide microarrays – pioneered by Affymetrix

On Affymetrix GeneChips, there are between 10 and 20 oligonucleotides for each gene. The choice of oligonucleotides is determined using a computer program that searches for nonoverlapping stretches of 20-25 bases that are unique in the genome (in order to prevent cross-hybridization). Furthermore, the computer searches for oligonucleotides that will fit empirically derived design rules that dictate the ratio of G–C pairs vs. A–T pairs and that attempt to reduce the likelihood that the oligonucleotide will hybridize to itself, creating hairpin structures.