WHAT IS THE PRINCIPLE OF HYB & SEQ?
The principle of Hyb & Seq is mismatch detection by enzymatic reaction performed on the microarray chip. A target DNA sequence is amplified with incorporation of a common tag antisense to the detection primer. A CustomArray 12K chip is synthesized with oligonucleotide probes that are complimentary to the target sequence and carry terminal mismatches. The amplified target sequence is hybridized to the CustomArray chip in the presence of the common labeled detection primer (Cy3-tag). Oligonucleotide probes of the chip and the detection primer anneal to the target. Subsequent incubation with enzymes results in extension of the detection primer and ligation to the probes, but only in the cases of exact match between the probe terminal nucleotide and the target sequence. After the enzymatic reactions are complete, DNA hybrids are denatured by stringent washing. The labeled detection primers are washed away from the chip except for those ligated to the probes. Therefore, signal is only detec
