Biomedical Engineering Reference
In-Depth Information
2.5
Applied Biosystems/SOLiD
The SOLiD system, developed in the laboratory of George Church, was acquired
by Applied Biosystems in 2006 (Fig.
1.1
)
. In essence, the sequencer uses short-
read sequencing technology by ligation (
com
). The resequencing of the
Escherichia coli
genome by SOLiD sequence was
reported in 2005 (Shendure et al.
2005
). In 2007, Applied Biosystems made
improvements to the technology and commercially released the SOLiD instrumen-
tation. Similar to 454 technology, SOLiD library DNA fragments are ligated to
oligonucleotide adapters, attached to beads, and clonally amplifi ed by emulsion
PCR. Then, beads are immobilized onto a derivatized-glass fl ow-cell surface, and
sequencing begins by annealing a primer oligonucleotide complementary to the
adapter at the adapter-template junction (Voelkerding et al.
2009
). The primer is
oriented to provide a 5
phosphate group for ligation to interrogation probes during
the fi rst “ligation sequencing” step. The 8-mer interrogation probe consists of (in
the 3
′
direction) two probe-specifi c bases followed by six degenerate bases
with one of four fl uorescent dyes at the 5
′
-to-5
′
end. The two probe-specifi c bases con-
sist of one of 16 possible two-base combinations. For the fi rst ligation sequencing
step, the 16 possible two-base interrogation combinations, and a thermostable
ligase are present. There is a probe competition for template sequence annealing
located adjacent to the primer. Post-annealing, ligation occurs and a wash to
remove unbound probe follows (Mardis
2008
). The fl uorescent signal is recorded
before cleavage of the ligated probes, a wash is performed to remove the fl uor, and
the 5
′
phosphate group is regenerated. In the next steps, ligation of interrogation
probes to the 5
′
phosphate group of the preceding 5-mer is performed. The fi rst
primer is extended by seven cycles of ligation, known as a round. Denaturation of
the synthesized strand is performed and a new sequencing primer offset by one
base in the adapter sequence (n1) is annealed. A total of fi ve rounds are performed,
each with a new successively offset primer, permitting each nucleotide to be
sequenced twice. The sequence can be obtained by examining the 16 two-base pos-
sible interrogation probes.
At the end of 2007, ABI released the fi rst SOLiD system with a read length of
35 bp and an output of 3 Gb per run. Capping, to decrease signal deterioration, cou-
pled with high-fi delity ligation chemistry and interrogation of each nucleotide base
twice during independent ligation cycles yields SOLiD can reach an accuracy of
99.9 % for a known target at a 15-fold sequence coverage over sequence reads of 25
nucleotides (Liu et al.
2012
). Five system upgrades followed in the next 3 years. In
2010, the SOLiD 5500xl sequencing system was released with improvements that
included improved read length, accuracy, and data output of 85 bp, 99.99 %, and
30 Gb per run, respectively. The shortcoming of this platform is the short-read length.
The applications of this platform include whole-genome resequencing, targeted rese-
quencing, transcriptome profi ling, and epigenome. Like other NGS systems,
SOLiD's computational infrastructure is expensive. A complete run is 7 days and
′
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