LI-COR sequencers can routinely provide 800-1100 nucleotide, one-pass reads with accuracy better than 99% on single and double-stranded DNA templates. These instruments produce long nucleotide reads through the use of a focusing microscope mounted on a scanning laser to detect near-infrared fluorescence labeled products from DNA sequencing reactions. The number of nucleotides determined from a set of four lanes is a function of the quality of DNA template and the length of the gel used to resolve the reaction products. The high accuracy levels are, in part, attributable to the use of the four-lane sequencing strategy. Since raw data are de-convoluted (fluorescent labeled reaction products for each of the dideoxy-nucleotide chain terminations are electrophoresed in separate lanes) spectral overlap leading to incorrect or ambiguous base calls does not occur. The four lane presentation format also permits the rapid review and editing of sequence reads; however, this format allows the analysis of fewer reactions on a single gel.
The LI-COR 4200 IR2 system can detect primer labeled extension products of sequencing reactions at two different wavelengths. Coupled with the development of the dyes IRD700 and the previously described IRD41, two DNA sequence ladders can be simultaneously electrophoresed and analyzed on the LI-COR 4200 IR2 in a single set of gel lanes. The two-dye system doubles the number of sequencing reactions that can be loaded. Since there is no spectral overlap between IRD700 and IRD41, the LI-COR 4200 IR2 system provides high quality data comparable to that of the LI-COR 4000 using IRD41-labeled primers.