Automated hybridization/imaging device for fluorescent multiplex DNA sequencing

Patent · Sun Jan 01 00:00:00 EST 1995

OSTI ID:870183

Weiss, Robert B ^[1]; Kimball, Alvin W ^[1]; Gesteland, Raymond F ^[1]; Ferguson, F Mark ^[1]; Dunn, Diane M ^[2]; Di Sera, Leonard J ^[1]; Cherry, Joshua L ^[1]

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1. Salt Lake City, UT
2. West Valley City, UT

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, then an 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.

• Research Organization: Univ. of Utah, Salt Lake City, UT (United States)
• DOE Contract Number: FG02-88ER60700
• Assignee: University of Utah (Salt Lake City, UT)
• Patent Number(s): US 5470710
• OSTI ID: 870183
• Country of Publication: United States
• Language: English

Related Subjects

automated
hybridization
imaging
device
fluorescent
multiplex
dna
sequencing
method
disclosed
integrated
chamber
comprises
mounting
membrane
containing
size-fractionated
reaction
products
apparatus
fluid
delivery
light
image
recording
fluorescence
emanating
programmable
controller
controlling
operation
hybridized
probe
enzyme
alkaline
phosphatase
bound
binding
moiety
fluorogenic
substrate
benzothiazole
derivative
introduced
converts
product
illuminated
beam
excites
produce
pattern
imaged
ccd
camera
component
obtain
series
digital
signals
converted
string
nucleotides
corresponding
nucleotide
sequence
reader
applicable
membrane-based
applications
colony
plaque
southern
northern
western
blots
digital signals
chamber device
ccd camera
digital signal
nucleotide sequence
imaging apparatus
imaging device
reaction product
reaction products
dna sequencing
multiplex sequencing
membrane containing
programmable control
fluorescent product
controller apparatus
sequencing reaction
nucleotides corresponding
programmable controller
alkaline ph
delivery apparatus
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