Automated hybridization/imaging device for fluorescent multiplex DNA sequencing

Patent · Sat Dec 31 23:00:00 EST 1994

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]

---

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: University of Utah
• 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

/435/
alkaline
alkaline ph
apparatus
applicable
applications
automated
beam
benzothiazole
binding
blots
bound
camera
ccd
ccd camera
chamber
chamber device
colony
component
comprises
containing
controller
controller apparatus
controlling
converted
converts
corresponding
delivery
delivery apparatus
derivative
device
digital
digital signal
digital signals
disclosed
dna
dna sequencing
emanating
enzyme
excites
fluid
fluorescence
fluorescent
fluorescent product
fluorogenic
hybridization
hybridized
illuminated
image
imaged
imaging
imaging apparatus
imaging device
integrated
introduced
light
membrane
membrane containing
membrane-based
method
moiety
mounting
multiplex
multiplex sequencing
northern
nucleotide
nucleotide sequence
nucleotides
nucleotides corresponding
obtain
operation
pattern
phosphatase
plaque
probe
produce
product
products
programmable
programmable control
programmable controller
reaction
reaction product
reaction products
reader
recording
sequence
sequencing
sequencing reaction
series
signals
size-fractionated
southern
string
substrate
western