Figure 5.Applied Biosystems (Life Technologies) SOLiD sequencing by ligation.

Top: SOLiD color-space coding. Each interrogation probe is an octamer, which consists of (3'-to-5' direction) 2 probe-specific bases followed by 6 degenerate bases (nnnzzz) with one of 4 fluorescent labels linked to the 5' end. The 2 probe-specific bases consist of one of 16 possible 2-base combinations. Bottom: (A), The P1 adapter and template with annealed primer (n) is interrogated by probes representing the 16 possible 2-base combinations. In this example, the 2 specific bases complementary to the template are AT. (B), After annealing and ligation of the probe, fluorescence is recorded before cleavage of the last 3 degenerate probe bases. The 5' end of the cleaved probe is phosphorylated (not shown) before the second sequencing step. (C), Annealing and ligation of the next probe. (D), Complete extension of primer (n) through the first round consisting of 7 cycles of ligation. (E), The product extended from primer (n) is denatured from the adapter/ template, and the second round of sequencing is performed with primer (n – 1). With the use of progressively offset primers, in this example (n – 1), adapter bases are sequenced, and this known sequence is used in conjunction with the color-space coding for determining the template sequence by deconvolution. In this technology, template bases are interrogated twice.

Republished from reference 5 with permission from the publisher.