Automated hybridization/imaging device for fluorescent multiplex DNA sequencing
A method is disclosed for automated multiplex sequencing of DNA with an integrated automated imaging hybridization chamber system. This system comprises an hybridization chamber device for mounting a membrane containing size-fractionated multiplex sequencing reaction products, apparatus for fluid delivery to the chamber device, imaging apparatus for light delivery to the membrane and image recording of fluorescence emanating from the membrane while in the chamber device, and programmable controller apparatus for controlling operation of the system. The multiplex reaction products are hybridized with a probe, the enzyme (such as alkaline phosphatase) is bound to a binding moiety on the probe, and a fluorogenic substrate (such as a benzothiazole derivative) is introduced into the chamber device by the fluid delivery apparatus. The enzyme converts the fluorogenic substrate into a fluorescent product which, when illuminated in the chamber device with a beam of light from the imaging apparatus, excites fluorescence of the fluorescent product to produce a pattern of hybridization. The pattern of hybridization is imaged by a CCD camera component of the imaging apparatus to obtain a series of digital signals. These signals are converted by the controller apparatus into a string of nucleotides corresponding to the nucleotide sequence an automated sequence reader. The method and apparatus are also applicable to other membrane-based applications such as colony and plaque hybridization and Southern, Northern, and Western blots. 9 figs.
- Research Organization:
- University of Utah
- DOE Contract Number:
- FG02-88ER60700
- Assignee:
- Univ. of Utah, Salt Lake City, UT (United States)
- Patent Number(s):
- US 5,470,710/A/
- Application Number:
- PAN: 8-141,234; CNN: Grant R01-HG00517-02; Grant P30-HG00199-03
- OSTI ID:
- 131929
- Country of Publication:
- United States
- Language:
- English
Similar Records
Enzyme-linked fluorescent detection for automated multiplex DNA sequencing
Multiplex sequencing and cosmid mapping: Progress report