Biology Reference
In-Depth Information
7.11.1.3 Applied Biosystems SOLiD Sequencer
Applied Biosystems SOLiD sequencing uses ligase to synthesize the DNA ampli-
cons on the surface of tiny 1-
μ
m paramagnetic beads (
Shendure et al. 2005
) and
has been commercially available since 2008. First, a library of the DNA is pre-
pared, and emulsion PCR is performed in microreactors (with template DNA,
PCR reaction components, beads, and primers). After the PCR, the templates are
denatured and the beads are separated that contain extended templates. The
template on the beads then undergoes a modification to attach it to the slide.
Sequencing is by ligation. Primers hybridize to the adapter sequence on the
beads. A set of four fluorescently labeled two-base probes compete for ligation
to the sequencing primer. Multiple cycles of ligation, detection, and cleavage
are performed with the number of cycles determining the read length. After
a series of cycles, the extension product is removed and the template is reset
with a primer complementary to the n-1 position for a second round of ligation
cycles. Five rounds of primer reset are completed for each sequence tag, so that
every base is evaluated in two independent ligation reactions by two different
primers. After ligation, the images are obtained in four channels, thus obtain-
ing data for the same base positions across all beads. Details of this process are
available at the Applied Biosystems website. This approach typically produces
reads of 35bp, and
Hayden (2009)
reported the SOLiD system could produce
1000 bp of sequence for US$0.002.
7.11.1.4 Standards
Field et al. (2009)
proposed that sharing of “Omics” data should be adopted as
a standard component of such research and that databases should be developed
to assist in this. Standard methods for describing, formatting, submitting, and
exchanging data need to be developed for the enormous amounts of genetic
data already developed and to be developed. Data release and exchange need
to become part of the scientific culture, even though such requirements add
costs to the project. Collecting, holding, and exchanging data requires long-term
funding and a well-designed system, which is not fully available at this time.
One possible solution to the problem of storing vast amounts of data involves
cloud computing (
Nature Biotechnology
2010
). There are at least a hundred
research centers around the world generating thousands of gigabases of DNA
sequence data every week. For example, a single Illumina machine can gener-
ate up to 90 billion bases per run, which results in large amounts of raw data
to analyze and store. Data storage is possible for large sequencing facilities, but
even their space is becoming an issue. For smaller laboratories with limited data-
storage capacity, the option may be to handle and store data by cloud computing.
