On a steeplechase track about half the width of a human hair, Cornell University researchers are racing individual DNA molecules to learn how they move through tiny spaces. One of the surprising results: Large DNA molecules squeeze through certain small spaces faster than small ones.
The research is aimed at better understanding the methods biologists use to decipher genetic information contained in a sample of DNA. The findings could help in the development of new DNA chips to speed and simplify such processes as DNA fingerprinting or the sequencing of bases in DNA samples.
In sequencing, for example, a sample of DNA is chopped into many small fragments that are forced through an organic gel that separates them by length as they move through the gel's maze of microscopic pores. Exactly what happens as the fragments move through the maze is difficult to measure because the pores vary randomly in size and shape.
In order to do controlled studies, Harold Craighead, Cornell professor of applied and engineering physics, and graduate students Jongyoon Han and Stephen Turner have used the Cornell Nanofabrication Facility to make microscopically small sieves with openings of controlled sizes. In an experiment reported in the Aug. 23, 1999, issue of Physical Review Letters, they describe the movement of DNA through a microscopic channel with a series of narrow constrictions.
Using the photolithography techniques originally created to make electronic devices on silicon chips, the researchers carved channels 30 microns wide with alternating deep and shallow sections. The deep sections are about 1 micron high. The shallow sections are about 90 nanometers high, small enough to form "traps" that slow the progress of DNA fragments. (A micron, or micrometer, is one-millionth of a meter; a nanometer is one-billionth of a meter, or about three times the diameter of an atom.)
DNA molecules in a water solution were introduced at one end of the channel, and an electric field was applied to pull them toward the other end. Even a large molecule like DNA is too small to see under a light microscope; to observe the behavior of individual molecules, the samples were tagged with fluorescent dyes, and researchers observed their movement through a glass plate covering the device. In video, individual molecules traveling through the channel appear as wiggling blobs of light moving across the screen.
Floating freely in water, a DNA chain contracts into a roughly spherical blob. In order to pass through the shallow traps, the molecules have to stretch out into a flatter shape. The time to pass through the channel varies with the time it takes individual molecules to deform and slip through the traps.
The researchers worked with two kinds of DNA. One, called T2 chains, are 4.3 times as long as the other, called T7 chains, and, consequently, contract into a much larger blob. Surprisingly, the larger molecules move through the channel faster. While both molecules move through the deep portions of the channel at the same speed, the smaller molecules wait longer at the beginning of each trap before squeezing through.
The reason, Craighead says, is that after a molecule presses against the start of a trap, a portion of the chain extends into the narrow space, and the rest of the molecule then stretches out to follow. The larger molecule, he explains, presses against the opening over a wider area, offering more places where a bit of itself can pull out to form such a "beachhead."
The researchers made channels with various spacings and found that the time taken by a molecule moving through the entire course also varies with the length of the deep sections of the channel. If the deep section is long enough, a molecule can return fully to its spherical shape before reaching the next trap. Otherwise the molecule might still be partly stretched when it reaches the next trap and can move into it more quickly.
"It was generally believed that it would be more difficult for longer DNA molecules to pass through the small constrictions," Craighead says, pointing out that in gel electrophoresis, longer molecules generally move slower than shorter ones. The opposite occurs in this device, he says, because of the deformability of the DNA polymers and the shape of the constriction. "It is important to recognize that a DNA molecule is not a solid particle and can deform to pass through the constriction," he says.
The paper, titled "Entropic Trapping and Escape of Long DNA molecules at Submicron Size Constriction," appears in the Aug. 23 issue of Physical Review Letters. The research was funded by the National Institutes of Health.