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Genomics
Our engineers and scientists are creating technologies that could revolutionize the field of genomics. We have built several models of a maskless array synthesizer, and we are applying that technology to a next generation system – the automated gene synthesizer – that will give individual scientists the means to build customized genes in their own labs in a matter of hours.
 Our understanding of genetics has increased tremendously and continues to develop at an accelerated pace. A mere 50 years after the double helix structure of DNA was discovered, we now have the complete genetic code of a person, all 3 billion base pairs comprising more than 20,000 genes.
The next step is to understand the role of each gene in a person’s life: Which genes predispose people to disease or cancer? What causes genes to be over or under expressed? Which genes are targeted by viruses and bacteria?
We hope to expedite the process by building upon existing DNA microarray technology. Also called gene chips, DNA microarrays are formed by using methods adapted from the semiconductor industry. They can hold up to hundreds of thousands of DNA strands on a coin-sized chip. Traditionally, the DNA is synthesized by using physical masks to selectively illuminate regions on the chip. The light removes a chemical protecting group and allows for the addition of the next DNA base (A, T, C or G) in that position. Each layer of the chip requires four physical masks, one for each DNA base. Therefore, a chip containing 25 base oligonucleotides requires 100 masks. [Read more about the process]
At a cost of thousands of dollars per mask, the technology is expensive, and any change to the chip calls for a new set of masks. To bypass this time consuming and costly step, researchers at the University of Wisconsin Center for NanoTechnology developed a maskless array synthesizer (MAS). The MAS adds nucleotides using the Affmetrix (GeneChip) process, but instead of controlling light with physical masks, light is selectively directed by an array of 786,000 micromirrors controlled by a computer chip. [Click here to see an animation.] The chip, developed by Texas Instruments, is the Digital Micromirror Device and is identical to those that control large digital television screens and projectors.
 A boon of the MAS technology is the automated synthesis of large DNA sequences. It has the potential to pack more DNA strands – up to an entire genome – on a single chip. Our current aim is to make customized strands of DNA, 10,000 base pairs long, on demand with a 24-hour turnaround. We plan to streamline the process into an automated, high throughput system.
To this end, we are developing software that would divide a DNA strand of 10,000 base pairs into unique oligonucleotides of approximately 40 base pairs in length. Each 40mer will be designed so that 20 base pairs overlap with a complementary sequence of an adjacent 40mer. The 40mers will be synthesized on the chip, then cleaved from the chip surface and amplified by PCR into DNA fragments of 500-1,000 base pairs. These medium-sized fragments will also incorporate overlapping regions, which are linked to form a DNA strand of 10,000 base pairs. [View image] To ensure a reliable gene product, each amplified section will be filtered with the bacterial protein MutS. MutS binds mistakes in DNA chains and removes errors.
 Our current process is only partially automated. Oligonucleotides are eluted as 40mers and synthesized off-chip using PCR. An automated process will not only reduce errors and expedite the synthesis, but will increase the concentration of the final product.
The automated gene synthesizer could promote a myriad of applications. Scientists could conveniently program the construction of any plant or animal gene, insert a bacterial sequence into a plant gene, create easy knock-in or knock-out genes, or compare the interaction of proteins or other biological structures among genes of different species. They could test a variety of induced errors and change sequences on a daily basis. Additional applications could include synthesizing proteins, detecting SNPs and constructing siRNA libraries.
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