US 5,288,644 AGranted

Instrument and method for the sequencing of genome

AssigneeThe Rockefeller University

InventorsRonald C. Beavis, Brian T. Chait

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Abstract

A method for DNA sequencing, and particularly for sequencing of the entire human genome. Different base-specific reactions are utilized to use different sets of DNA fragments from a piece of DNA of unknown sequence. Each of the different sets of DNA fragments has a common origin and terminates at a particular base along the unknown sequence. The molecular weight of the DNA fragments in each of the different sets is detected by a matrix assisted laser absorption mass spectrometer to determine the sequence of the different bases in the DNA. The methods and apparatus of the present invention provide a relatively simple and low cost technique which may be automated to sequence thousands of gene bases per hour, and eliminates the tedious and time consuming gel electrophoresis separation technique conventionally used to determine the masses of DNA fragments.

Claims8

Dependent claims

A method of DNA sequencing of the genome by detecting the mass weights of DNA fragments comprising the steps of:

(a) producing a piece of DNA of unknown sequence;
(b) performing four different base-specific reactions on the piece of DNA to produce four different DNA sets containing DNA fragments each DNA set having a common origin and terminating at a particular base along the unknown sequence;
(c) selecting a solid matrix having a strong absorption band at the wavelength of light produced by a laser and placing one of the DNA sets on the matrix, the strong absorption band of the matrix having a longer wavelength than the absorption bands for the DNA set;
(d) without separation of the DNA fragments by electrophoresis or other separation methods and without labeling the DNA fragments with radioactive, fluorescent or other labeling means, striking the DNA fragments on the solid matrix within one of the DNA sets with a series of laser pulses from the laser to desorb ions of the DNA fragments and produce ionized DNA fragments;
(e) detecting the mass weights of the ionized DNA fragments by a time of flight mass spectrometer;
and
(f) repeating steps (d) and (e) for the other DNA sets to determine the sequence of bases in the DNA.

4 more elements

Dependencies are inferred from the claim text.

Facts

Application: 07/957,688Filed 13 Nov 1992 · continuation
Priority: 4 Apr 1990
Granted: 22 Feb 1994
Term ends: Not estimatedThe source records don’t support an estimate.

Classification25 codes

H01J 49/0413
Sample holders or containers: for automated handling
B01J 19/0046
Chemical, physical or physico-chemical processes in general; Their relevant apparatus: sequential or parallel reactions, e.g. for the synthesis of…
C12Q 1/6872
Methods for sequencing: involving mass spectrometry
H01J 49/164
Using photoionisation, e.g. by laser: laser desorption/ionisation, e.g. matrix-assisted laser desorption/ionisation [MALDI]