Sequence Sudoku

May 15, 2007 | The basis of Applied Biosystem’s new SOLiD platform differs from traditional Sanger sequencing in several respects. First and foremost, it uses DNA ligases to anneal fluorescently tagged DNA cassettes to the template, rather than polymerases. But the ingenious twist is the way that sequence is actually interrogated and read out.

Each SOLiD probe interrogates two adjacent bases at a time, not a single base. But as it uses only four fluorescent tags, each color tag matches four possible dinucleotide pairings (see panel A). For example, blue corresponds to AA, CC, TT, or GG. Thus, any color by itself cannot reveal the identity of a particular base. But once the identity of the first base is known, then the rest fall neatly into position.

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For example, if the first base is an A, and the first color is blue, then the first two bases must be A-A (because the other possibilities are by definition excluded). Now lets turn to the second color — green. Since we just deduced that the second base in the sequence is an A, we deduce the identity of the third base by reading out which green dinucleotide pair starts with an A. Thus, the third base is C. This process continues for the read length of 25 bases (see panel B).


Although the individual read lengths are very small, the system offers some significant advantages. The system neatly distinguishes between naturally occurring variants (SNPs) and random sequencing errors. In most sequencing systems, there is no way to discern a genuine SNP versus a sequence error. But in SOLiD, a random sequence error will only change a single colored tag. A genuine SNP, by contrast, is revealed as changes in consecutive color tags, because the altered base occurs in two successive nucleotide pairings.  -- K.D.

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