[tt] [Fwd: [wta-talk] DNA in Real Time]

[Brian Atkins]

-------- Original Message --------
Subject: [wta-talk] DNA in Real Time
Date: Tue, 16 Sep 2008 19:41:22 +0300
From: arsen zahray <menkaur at gmail.com>
Reply-To: World Transhumanist Association Discussion List 
<wta-talk at transhumanism.org>
To: <wta-talk at transhumanism.org>

>From http://www.technologyreview.com/Biztech/21376/page1/ 



Pacific Biosciences aims to bring sequencing technology to the doctor's
office.



In a nondescript office park in the northeast side of Menlo Park, CA, the
next genomics revolution may be taking place. There, 12 prototypes of a new
sequencing machine developed by startup Pacific Biosciences are churning out
reams of DNA sequence as fast as built-in cameras can record it. The deep
freezer-size boxes, covered for the time being in red plastic sheeting, are
performing a novel feat: reading single strands of DNA in real time. The
machine's creators hope that innovation will result in a process that
operates fast and cheaply enough to make sequencing a routine component of
medical care.



The company, founded in 2004, made a splashy debut almost a year ago,
showcasing its technology to the scientific community for the first time at
a sequencing conference in Florida. Steve Turner, the company's founder and
chief technology officer, wowed the audience with a video of single molecule
sequencing in progress, the product of a proof-of-principle experiment
reading a 150-base-pair length of DNA. Since then, the startup has garnered
$100 million more in funding--for a total of $178 million to date.



Pacific Biosciences' central innovation is a small chip composed of a
100-nanometer-thick metal film deposited on a silicon-dioxide substrate and
dotted with thousands of tiny wells, each only tens of nanometers in
diameter. Before sequencing begins, an enzyme called DNA polymerase is
immobilized at the bottom of the well, along with the strand of DNA to be
read. Fluorescently labeled bases--each of the four DNA letters labeled with
a different marker--are then flooded onto the plate, randomly diffusing into
each well. When the correct base diffuses into the bottom of a well, the
enzyme attaches it to the growing strand of DNA.



The wells are so small that fluorescent light shined through the bottom of
the plate penetrates only the lower 20 to 30 nanometers of each well,
meaning that only the bases being actively attached to the DNA molecule
light up. That allows a camera to capture the sequencing reaction as it
happens, leaving any irrelevant chemical activity in the dark. "The
waveguide is the first technology to allow observation of the polymerase in
action at physiologically relevant concentrations," says Turner.



In a video showing the sequencing reactions in action, a series of lights
scattered across the screen burst and fade in quick succession. (A computer
detects which base is added with each burst by its position within a well.)
The machines can currently sequence 12 million bases of DNA per hour, about
one-third of a percent of a human genome. The video must actually be slowed
to be viewed: the reactions occur too fast to be visible to the human eye.



Despite its early success, it's not yet clear whether the company's
innovative approach will surpass "next generation" sequencing technologies
already in use. Pacific Biosciences plans to release a commercial product in
2010 and will announce the target sequencing capacity for that machine early
next year. New types of sequencing machines, such as those developed by
Roche Applied Sciences and Illumina, have already revolutionized genomics,
allowing scientists to sequence huge volumes of DNA and reportedly dropping
the cost of a human genome to less than $100,000.



But Turner is confident that his method has advantages, especially in the
clinical diagnostics market. The most advanced sequencing approaches
currently on the market stop the sequencing reaction after the addition of
each base, wash away extra bases, snap a picture, and then repeat. Real-time
sequencing is faster and uses fewer chemicals, making it much cheaper. "In
the long run, the reagent cost dominates in sequencing," says Chad Nusbaum,
codirector of the Genome Sequencing and Analysis program at the Broad
Institute, in Cambridge, MA.

Both Roche and Illumina have ramped up speed by running a massive number of
sequencing reactions in parallel. However, these methods generate relatively
short lengths of sequence, about 50 to 200 bases pairs, which must then be
computationally assembled into a complete sequence. The shorter the piece,
the more computationally difficult it is to sew them together. "With a 35
base read, you can't assemble 25 percent of the genome," says Turner.



Pacific Biosciences has already generated what Turner believes to be "the
longest sequencing trace ever made." In a proof-of-principle experiment,
scientists continuously sequenced the same 135 base circle of DNA 12
times--it was like repeatedly driving around a rotary. While the company
hasn't yet repeated the feat with natural DNA, the ability to generate long
reads could be especially important when sequencing unknown DNA or stretches
with a highly repetitive series of bases. "Many of the dynamic regions of
the genome that are associated with disease consist of duplicated and
complex repetitive sequences that cannot be accurately assessed at present,"
says Evan Eichler, a geneticist at the University of Washington, in Seattle.
"Long-sequence reads are necessary to comprehensively understand human
genetic variation."



Pacific Biosciences' machine may also enable scientists to generate more
accurate sequences. Existing methods generate a consensus sequence by
reading the same section of multiple copies of DNA, which may have some
copying errors, and pooling the results. "All short-read technologies are
less accurate than traditional methods," says Nusbaum. With Pacific
Biosciences technology, scientists would theoretically be able to sequence
the same piece of DNA multiple times. "That means we'll be able to detect
rare mutations with unprecedented accuracy, orders of magnitude better than
others," says Turner.



He adds that the long reads, high accuracy, and quick turnaround time make
the technology ideal for diagnostics, such as those for cancer, which
involves long stretches of repetitive sequences, and infectious disease, in
which small sequence changes may drastically change the infectiousness and
virulence of a pathogen.



Perhaps the biggest excitement comes from Turner's predictions for the
future. He says that it will be easy to further improve the company's
technology with higher-resolution cameras, faster DNA polymerases, and more
densely packed chips. Currently, each chip has thousands of wells, only
about one-third of which are used. (The remaining two-thirds house either no
enzyme molecules or more than one, and thus fail to generate useful
information.) But the chips, which are created using semiconductor
fabrication methods, have the capacity to hold 10 million wells.



Turner calculates that with a camera that can track one million wells, a
polymerase that operates at about 50 bases per hour (the current rate is
10), and full use of all the wells on the plate, Pacific Biosciences
technology could read 100 gigabases an hour. That translates to full
coverage of a human genome--the same genome sequenced 15 times--in just 15
minutes.



"I'm pretty excited about the possibilities of this technology, but I remain
to be convinced," says Nusbaum. "They still have some significant technical
hurdles ahead of them."



-- 
Brian Atkins
Singularity Institute for Artificial Intelligence
http://www.singinst.org/
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