Gene chip engineers
Peter Hoyt checks the operation of the Packard multiprobe liquid-handling system and mechanical gripper. This state-of-the-art robotic tool is used to prepare samples (e.g., cloned DNA) for use on microarrays to analyze gene expression.
What does a gene do in a mouse, fish, or some other organism? One technology that
allows biologists to spy on gene activity is the microarray, a microscope glass slide
dotted with an orderly array of DNA sequences.
Mitch Doktycz, Peter Hoyt, and their colleagues in the Life Sciences Division (LSD)
specialize in designing and using microarrays, or gene chips, to help determine which
genes are expressed as a result of specific diseases or exposures to environmental
toxins. They also have developed technologies to speed up and reduce the costs of
preparing DNA probes for gene chips, as well as genetic material from mice, fish, and
"We look at hundreds to thousands of
samples of biological liquids
simultaneously on our microarrays,"
Doktycz says. "We are evaluating
expressed messenger RNA (mRNA)
isolated from various mouse tissues,
including skin, liver, lung, brain,
muscle, kidney, fat, heart, pancreas,
spleen, gut, and testes.
In collaboration with Ed Michaud's
group in LSD (see Complex
Biological Systems in Mice),
Doktycz's group has analyzed gene expression in samples collected from mice afflicted
with skin disease to figure out which genes are altered in the diseased mice compared
with equivalent genes in normal mice. A gene that is altered produces abnormally high
or low levels of mRNA, which eventually results in altered levels of protein.
Doktycz is also collaborating with ORNL's Russ Knapp and Ed Michaud to determine
the genes that are altered in mouse models of obesity and diabetes. Specifically, these
experiments use gene chips to determine how new anti-diabetic drugs treat the disease
and affect expression patterns of these genes.
Working with Mark Greeley of ORNL's Environmental Sciences Division (ESD),
Doktycz and his colleagues have developed a "zebrafish tox-chip microarray." It is used
to determine which genes are turned on in zebrafish embryos exposed to
Besides applying microarrays to gene expression and genome studies, these ORNL
researchers are developing more economical ways to prepare samples—extracting
mRNA from cells for gene expression studies and attaching various DNA probes to
"When we complete development of automated sample processing, hundreds of tissue
samples will be processed in an afternoon," Doktycz says. "Using conventional
techniques, it can take three to four days to analyze 12 samples. With our method, 96
samples can be prepared in parallel in just a few hours. This sample productivity is
needed because we can make more than 100 gene chips in less than a day. Currently,
we can print several thousand DNA spots on each gene chip."
Hoyt has developed an inexpensive, high-throughput liquid-handling method of
extracting mRNA from tissues. A snippet of mouse skin or other tissue is broken up into
cells by simultaneously homogenizing the samples. After the cells are placed in a
microtiter plate, mRNA is isolated from them using an automated procedure. Hoyt is
now beta testing a new Packard Bioscience robotic instrument for "walk-away"
automated processing of mRNA samples. The same instrument is used to prepare the
complementary DNA test samples placed on the gene chip (where they will pair with
the matching mRNA samples).
Doktyz has been making a mark in
the field of spotting technologies. He
recently helped devise a commercial
inkjet technology for dispensing
microscopic drops of biological fluids
at high speeds, a technology that
could hasten the development of new
therapeutic drugs. He worked on this
project with Rheodyne, a California
company that makes high-end valves,
under a cooperative research and
development agreement. The
resulting "hybrid valve" is now
produced commercially by
Innovadyne Technologies, Inc., a
Rheodyne spin-off company.
Gene chips and related technologies are revolutionizing biological studies. To help meet
the need for faster, better, and cheaper ways to spy on genes, Doktycz and his
colleagues are being ingenious.