Electrophoresis and fluorescence detection method

Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid

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2041828, 204299R, 36441301, 422 681, 435912, C12Q 168, C25B 100, B01D 6142, G01N 1506

Patent

active

057861420

ABSTRACT:
An improved electrophoresis and fluorescence detection method for nucleotide sequences comprises a fluorescence sensing region along the path of nucleotide detection, coupled with amplification and integration in an integrator of output signals in the form of activity peaks. The output signal, which is converted to a voltage signal, is summed with a programmable offset generated by an inexpensive eight-bit D/A converter. The offset signal is selected to establish a lower starting point for the dynamic range of analog-digital conversion, and is selected to null some or all of the background fluorescence level. The integrator is switchable under program control. The integrator is switched on for long and short integration intervals. The short intervals permit sensing over a dynamic range accommodating very high levels of fluorescence; very high peaks may be measured and features of the peaks distinguished. The long intervals permit sensing over a dynamic range that is optimized for the peaks associated with the smaller peaks of individual nucleotides. In this way, the dynamic range of the analog-digital conversion permits the highest possible resolution over the range of interest during the time in which the sequencing of the nucleotides takes place. The method of nucleotide sequencing and analysis is fast, economical, and yields data with high resolution.

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Swerdlow et al, "Reloading and stability of polyacrylamide slab gels for automated DNA sequencing", Biotechniques 16(4):684-693, 1994.

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