Applied Biosystems SOLiD™ 4 System SETS Software User Guide ...

Applied Biosystems SOLiD 4 System SETS Software User Guide
Appendix B Advanced Topic: Data Analysis Overview
Fundamentals of color-space analysis B
Figure 3 Requirements that assign color for a 2-base code
Principles of ligation-based chemistry and 2-base encoding
The SOLiD 4 System’s sequencing technology is based on
sequential ligation of dye-labeled oligonucleotide probes. Each
probe assays two base positions at a time (Figure 2). The system uses
four fluorescent dyes to encode for the sixteen possible 2-base
combinations. Multiple ligation cycles of probe hybridization,
ligation imaging, and analysis are performed to extend the strand
from a primer hybridized to a ligated adaptor by the immobilized
bead (P1 adaptor). The resulting product is then removed and the
process repeated for 5 more rounds with primers hybridized to
positions n-1, n-2, and so on, in the P1 adaptor.
There are several fundamental properties unique to ligation-based
sequencing. These properties contribute to the high accuracy
inherent in the SOLiD 4 System. The advantages of these
properties and their contribution to data quality are:
Two bases are interrogated in each ligation reaction, increasing
specificity.
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