US 5,514,256 AGranted

Apparatus for improved DNA sequencing

AssigneeBattelle Memorial Institute

InventorsRichard J. Douthart, Shannon L. Crowell

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Abstract

This invention is a means for the rapid sequencing of DNA samples. More specifically, it consists of a new design direct blotting electrophoresis unit. The DNA sequence is deposited on a membrane attached to a rotating drum. Initial data compaction is facilitated by the use of a machined multi-channeled plate called a ribbon channel plate. Each channel is an isolated mini gel system much like a gel filled capillary. The system as a whole, however, is in a slab gel like format with the advantages of uniformity and easy reusability. The system can be used in different embodiments. The drum system is unique in that after deposition the drum rotates the deposited DNA into a large non-buffer open space where processing and detection can occur. The drum can also be removed in toto to special workstations for downstream processing, multiplexing and detection.

Claims16

Dependent claims

An apparatus for the sequencing of a DNA sample comprising:

(a) a gel support unit having
(1) an upper buffer chamber having an access slot, said upper buffer chamber mounted on,
(2) a plate assembly having cover plate and ribbon channel plate defining a plurality of sample loading spaces wherein each sample loading space is open to said access slot and adjacent to a polymerized gel within the ribbon channel plate, said plate assembly further having an interface edge placed adjacent to;
(b) a drum assembly having
(1) a drum with a deposition membrane mounted adjacent to and spaced apart from said interface edge, defining a plate-drum gap,
(2) a lower buffer chamber with a drum support unit, said drum placed upon said drum support unit and extending into said lower buffer chamber;
(c) an electrical circuit having
(1) an upper electrode mounted within said upper buffer chamber, and attached to a high voltage power supply,
(2) a lower electrode mounted within said lower buffer chamber and attached to said high voltage power supply, and
(3) a buffer solution(s) within said upper buffer chamber and said lower buffer chamber wherein a circuit is formed between said upper electrode and said lower electrode through said buffer solution in said upper buffer chamber, through said sample loading spaces, through said polymerized gel, and through a plate-drum interface defined in said plate-drum gap with said buffer solution from said lower buffer chamber, through buffer solution coating said deposition membrane, through the buffer solution within the lower buffer chamber to said lower electrode, and thence back to said power source;
wherein when said DNA sample is placed in the sample receiving wells it is electrophoretically separated in temporal distribution along said deposition membrane surface.

8 more elements

A method for sequencing DNA samples, comprising the steps of:

(a) providing a gel support unit having
(1) an upper buffer chamber having an access slot, said upper buffer chamber mounted on,
(2) a plate assembly having cover plate and ribbon channel plate defining a plurality of sample loading spaces wherein each sample loading space is open to said access slot and adjacent to a polymerized gel within the ribbon channel plate, said plate assembly further having an interface edge;
(b) placing a drum assembly adjacent to said gel said interface edge, said drum assembly having
(1) a drum with a deposition membrane mounted adjacent to and spaced apart from said interface edge, defining a plate-drum gap,
(2) a lower buffer chamber with a drum support unit, said drum placed upon said drum support unit and extending into said lower buffer chamber;
(c) providing electrical circuit having
(1) an upper electrode mounted within said upper buffer chamber, and attached to a high voltage power supply, and
(2) a lower electrode mounted within said lower buffer chamber and attached to said high voltage power supply,
(d) filling said sample spaces with said DNA samples;
(e) filling said upper buffer chamber and said lower buffer chamber with at least one buffer solution;
(f) rotating said drum and forming a plate-drum interface in said plate-drum gap with said buffer solution from said lower buffer chamber;
(g) imposing an electrical potential across the upper electrode and the lower electrode and passing electrical current between said upper electrode and said lower electrode through said buffer solution in said upper buffer chamber, through said DNA sample, through said polymerized gel, and through buffer solution coating said deposition membrane, through the buffer solution within the lower buffer chamber to said lower electrode, and thence back to said power source;
and
(h) electrophoretically separating said DNA sample in temporal distribution on said deposition membrane.

12 more elements

Dependencies are inferred from the claim text.