Rapid, Low-Cost DNA Testing

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Rapid, Low-Cost DNA Testing

http://www.medicalnewstoday.com/medicalnews.php?newsid=60336

Professor Rothberg of the University of Rochester Chemistry

Department received a NYSTAR grant in August 2006 to continue

working on a recent discovery by Huixiang Li, a research associate

in his group: how to rapidly test DNA to improve our health and make

sure we're drinking clean water and eating uncontaminated food. In

fact, his new method can be used to help forensics labs identify

criminals, test ponds and pools before children swim in them, and

identify harmful genetic sequences in medical research, to name only

a few applications. Rothberg's innovative procedure quickly and

inexpensively identifies genetic sequences in any sample of DNA.

The technology is a novel fluorescent DNA screening assay, which

rapidly determines whether specific DNA target sequences are present

in an analyte. In simple terms, the analyte contains the DNA target

sequences as well as other DNA sequences, and the assay filters out

only the targets. Professor Rothberg's assay is based on the

electrostatic properties of DNA.

The principle underlying the method is that single-stranded DNA and

double-stranded DNA have significantly different affinities for

attaching to ionically charged gold nanoparticles. Because ions have

electric charges, having gained or lost electrons, they attract

their opposites. An anion with a negative electric charge will

attract positive charges, a cation with a positive charge will

attract negative charges. Single-stranded DNA adsorbs on negatively

charged citrate ions on the gold nanoparticles while double-stranded

DNA does not. Given that both single-stranded and double-stranded

DNA are (nominally) negatively charged, this proven phenomenon

intrigues the research group.

The new assay determines whether a fluorescently-tagged short probe

sequence of single-stranded DNA matches a sequence in the target

analyte. When it does not, the fluorescently tagged probe adsorbs on

a gold nanoparticle and its fluorescence is quenched. If the probe

sequence is able to hybridize to the target, it will not adsorb on

the gold and its fluorescence persists.

The most widespread and common method of screening DNA is called gel

electrophoresis. Each test takes 1 hour and can cost as much as

$1.00. Setting up a lab for gel electrophoresis requires a capital

expenditure of $5,000. By contrast, Professor Rothberg's technique

only requires 5 minutes, and it costs approximately $0.05 (literally

five cents) per test. The capital expenditure to set up a lab with

the new technique is only $600.

Here's how Professor Rothberg's procedure is done:

Step 1. Hybridization. This takes 10 seconds and costs $0.025.

Step 2. Add gold colloid to the hybridization solution. This takes

10 seconds and costs $0.02.

Step 3. Add salt to the solution. This takes 10 seconds and costs

$0.01.

Step 4. Measure photoluminescence. This takes 1 minute.

It's as simple as that, yet nobody's ever done it before. The method

is so new that the University of Rochester filed patents for it in

2004 and 2006. In May 2005, Professor Rothberg created a company

called Diffinity Genomics, Inc. with two partners to further study

and commercialize his technique.

Professor Rothberg's method is part of a much larger process that

analyzes DNA. First, a technician extracts the DNA from the blood,

tissue, or food. This typically take up to an hour. Second, there is

generally not enough DNA to analyze, so it must be chemically

amplified. This also takes apprximately one hour. The new process

comes after these two steps, saving a final hour of work for the

technician, who ordinarily would be doing gel electrophoresis.

Perhaps more important than the savings in time and money, the new

method works to determine single-base mutations in DNA, whereas gels

cannot do this without even further processing. Professor Rothbergs

concludes, " This could be very important for applications in

personalized medicine where a particular DNA sequence will be linked

to a prescribed therapy. In fact, we see this happening already. "

For further details, see:

* Li, H., Rothberg, L.J., " Label-free colorimetric detection of

specific sequences in genomic DNA amplified by polymerase chain

reaction, " J. Am. Chem. Soc. 2004, 126, 10958-10961