Icahn School of Medicine at Mount Sinai
Mount Sinai - Twitter Mount Sinai - YouTube Mount Sinai - Facebook

Resources

Genomics Core Facility Microarrays BeadArray Technology

BeadArray Technology


Micrometer diameter beads positioned in etched silica holes

The Illumina approach for gene expression arrays is as follows: an oligonucleotide targeting a single gene transcript is synthesized and conjugated to a micrometer diameter silica bead. In addition an oligonucleotide with a unique sequence (molecular barcode) is also conjugated to the same bead. Millions of beads for thousands of gene targets are synthesized in parallel and then mixed together in equal proportion. Separately, glass slides with silica coating are manufactured and etched with an array of millions of small holes to fit the beads. Beads are spread onto the slides, enter the holes and are held by van der waals forces. The beads are positioned randomly, and each gene is targeted by multiple beads. In gene expression experiments, a fluorescently labeled RNA transcript is then hybridized to the array and signal read by an array scanner.

To find the bead to gene correspondence, a clever decoding strategy is used. All beads have a unique barcode, but there are subsequences that are shared between each. One oligonucleotide conjugated to a green dye, and another conjugated to a red dye are hybridized to the array and signal is read. Roughly half of the beads will have signal with green the other half with red. The array is subjected to denaturing conditions to remove the annealed oligonucleotides and the process is repeated with a different pair of green and red strands. Again, half of the beads will have green or red signal, but the signal may not the same as the previous iteration. For example, a bead may have four signal sequences: red-green, green-red, red-red or green-green. This process divides all beads into four groups. Using successive hybridization cycles, millions of beads can be individually decoded to identify which gene transcript they target. The same method applies for decoding beads in genotyping or methylation assays.