High-throughput DNA purification
|Automation is the buzz thats busting bottlenecks in large-scale population studies
With so much genomics research going on in the United States, it is easy to miss the hum of initiatives around the world. Researchers eager to lay hands on genes controlling disease have launched genomics projects in lands as distant and different as Iceland, Estonia, Sardinia, Newfoundland, and the South Pacific island kingdom of Tonga. Together, these population projects account for millions of large-volume patient samples, yet the idea of purifying DNA from even a few thousand samples remains a daunting prospect.
Scientists want to use automated genotyping to examine thousands of gene variants in millions of people, tracking down single genes controlling the rarest of diseases and locating the many genes that subtly influence our most widespread and complex diseases. Genomics in a faraway place can make those tasks easier: Quirks in isolated gene pools sometimes give researchers piercing insight into the genetic aspects of complex diseases. Genomic research in Icelandic populations has already resulted in the identification of genes linked to obesity, anxiety, diabetes, schizophrenia, Parkinsons disease, and rheumatoid arthritis.
Genomics is also spreading for other reasons. Icelands genomics project is aided by family trees that trace as far back as the 9th century, when ties of blood were recorded with the ink of blood, and animal skins served in place of paper. Estonia drives its project forward on sheer ambition. Its goal is to own the worlds largest genome project within five years, holding DNA samples from 1 million people2 of every 3 Estonians.
The buzz behind this genetic land rush is that population geneticists must find faster methods than preparing human genomic DNA by hand. Think of it this way. Which of the following phrases sounds out of place in a genomics laboratory?
The fact is that population genomics requires automation wherever possible, including DNA purification. Manual preps are fine for small projects, but competing in genomics by using hand-prepped DNA is like filling the tank at a NASCAR pit stop by using a teaspoon.
First to Finnish
Perola was given a thesis project on the genetics of hypertension. The complexity of the disease required DNA samples from many people, and he quickly became expert at isolating and purifying genomic DNA. Around the same time, epidemiologists at the institute began studies calling for enormous numbers of DNA samples. Perola was enlisted to help them. Soon, he found himself acting as the head of a team of eight, preparing DNA from 60,000 Finns entirely by hand, using the old phenolchloroform method (see box, DNA preparation: Old and new).
After receiving his doctorate, Perola followed Peltonen to her new laboratory in California, where he began using DNA microarrays to study the genetics of psychiatric and cardiovascular problems. Soon after his arrival, UCLAs human genetics department asked him to advise it on the best way to handle DNA preparation. He never doubted that the answer was to automate. The question was how.
The standard source of human DNA is blood drawn in 10-mL aliquots. Perola needed an instrument that would work with minimal human intervention, extracting DNA pure enough to store for years without degradation. Ideally, the preparation chemistry would not generate toxic wastes. He decided on the Autopure LS, introduced in 2000 by Gentra Systems, Inc. (Minneapolis; www.gentra.com).
DNA preps degrade for three reasons. They can be contaminated by bacteria, but storage at 4 ºC or lower prevents this, says Perola. More often, degradation is due to contamination of the sample with DNase from skin. Wearing gloves while preparing DNA can solve this. Generally overlooked, however, is degradation caused by continual cycles of freezing and thawing. Perola minimizes the problem by planning ahead.
We have a one-usage tube, as we call it, for aliquoting, he says, and a stock tube that is used once every few years. As an extra precaution, each sample has two stock tubes, which he stores at 20 ºC. A better temperature would be 80 ºC, which avoids slow evaporation and changes in DNA concentration, but freezing at 20 ºC is less expensive.
Perola strongly advises preparing DNA as soon as possible after drawing blood. Storing blood first and preparing DNA later sounds convenient but creates unnecessary work. His experience is consistent with findings that Gentra has published for some years: Storing blood results in lower yields (often substantially lower), no matter what the purification method.
What Perola did for UCLA, he hopes to repeat in his laboratory back home. He sees the difference that automation makes. At UCLA, one person can do about all that needs to be done, says Perola. The tech puts the samples on and comes back later. Theres the DNA. Another bonus is not having to worry about phenol and chloroform. This is important for us because both phenol and chloroform are carcinogenic and toxic, he says. And they also break DNA.
Thats an example of the kind of treasure you have once you have these materials, Perola says with enthusiasm. One analysis is not enough. You have to reanalyze in order to get all the information out. The discovery confirmed by their return to the archivesthat a gene on chromosome 7 is linked to heightwas published in the July 2000 issue of the American Journal of Human Genetics.
With the help of heavy-duty nucleic acid purification systems that can produce archival-quality DNA, investigators like Markus Perola can look forward to more successes. Each one is a part of the quiet hum of progress in international research, which is punctuated occasionally by the sound of a geneticist busting through another bottleneck. And in genomics research, busting barriers is the buzz.
Darin OBrien is a product manager at Gentra Systems, Inc. Send your comments or questions regarding this article to firstname.lastname@example.org or the Editorial Office by fax at 202-776-8166 or by post at 1155 16th Street, NW; Washington, DC 20036.