Apparatus and method for automated purification of extra-chromo-somal DNA from a cell suspension in a container

Claim 1

Method for automatic separation and recovery of extra-chromosomal DNA from a bacterial cell suspension, comprising introducing the bacterial cell suspension into a chamber, introducing a lysing solution into the chamber and mixing it with the contents thereof to lyse the cells therein, introducing a precipitating solution into the chamber and mixing it with the contents thereof to precipitate chromosomal DNA, proteins and cell debris therein, filtering the liquid contents of the chamber through a filter for removing precipitated protein, chromosomal DNA and cellular debris and feeding the filtered liquid to a collecting device, recovering extra-chromosomal DNA from the contents of the collecting device in the substantial absence of precipitated protein, chromosomal DNA and cellular debris, introducing a dissolving solution into the chamber and mixing it with the contents thereof to dissolve the precipitate remaining therein, and feeding the dissolved precipitate from the chamber to waste.

Claim 5

An apparatus for separating and recovering plasmid DNA from bacterial cells which contain plasmid DNA and chromosomal DNA; said apparatus comprising: a) a mixing chamber having a plurality of inlet and outlet ports for the passage of fluid therethrough and means for controlling the temperature of said chamber; b) an agitator in said mixing chamber; c) a first multi-port set of valves having a plurality of inlet and outlet ports; d) a lysing solution container and a conduit which connects said lysing solution container to an inlet port of said first multi-port set of valves for the passage of lysing solution therethrough; e) a precipitating solution container and a conduit which connects said precipitating solution container to an inlet port of said first multi-port set of valves for the passage of precipitating solution therethrough; f) a vacuum pump which is connected to an outlet port of said first multi-port set of valves; g) a bacterial cell suspension container and a conduit which connects said bacterial cell suspension container to an inlet port of said first multi-port set of valves for the passage of bacterial cell suspension therethrough; h) a conduit which connects an outlet port of said first multi-port set of valves to an inlet port of said mixing chamber for the passage of lysing solution, precipitating solution and bacterial cell suspension into said mixing chamber; i) a waste conduit connected to an outlet port of said first multi-port set of valves for the elimination of waste therefrom; j) a second multi-port set of valves having a plurality of inlet and outlet ports for the passage of fluid therethrough; k) a conduit which connects an outlet port of said mixing chamber to an inlet port of said second multi-port set of valves for the passage of fluid therethrough and means for filtering precipitated chromosomal DNA, protein and cellular debris from said fluid which passes therethrough for the production of a filtered fluid which contains dissolved plasmid DNA in the substantial absence of chromosomal DNA, protein and cellular debris; l) a sample collector for collecting the filtered fluid containing the dissolved plasmid DNA; said sample collector having an inlet and an outlet, set inlet of the collector being connected to an outlet port of said second multi-port set of valves for receiving said filtered fluid and said outlet of the colletor being connected to an inlet port of said second multi-port set of valves for returning said filtered fluid to said second multi-port set of valves; m) means for recovering the plasmid DNA, said means for recovering the plasmid DNA being connected by a conduit to an outlet port of said second multi-port set of valves for the passage of filtered fluid from the sample collector to the plasmid DNA recovery means via the second multi-port set of valves; n) a dissolving and cleaning solution container and a conduit which connects said dissolving and cleaning solution container to an inlet port of said second multi-port set of valves for the passage of dissolving and cleaning solution therethrough; o) a conduit which connects the second multi-port set of valves with the mixing chamber for passing the dissolving and cleaning solution from the second multi-port set of valves through said filter means and into the mixing chamber; and p) a conduit which connects an outlet port of said mixing chamber to an inlet port of said first multi-port set of valves for drawing bacterial cell suspension into said mixing chamber by action of said vacuum pump and for the elimination of the dissolving and cleaning solution through the waste conduit via the first multi-port set of valves.

Claim 6

An apparatus for separating and recovering plasmid DNA from bacterial cells which contain plasmid DNA and chromosomal DNA; said apparatus comprising: a) a mixing chamber having a plurality of inlet and outlet ports for the passage of fluid therethrough and means for controlling the temperature of said chamber; b) an agitator in said mixing chamber; c) a first multi-port set of valves having a plurality of inlet and outlet ports; d) a lysing solution container and a conduit which connects said lysing solution container to an inlet port of said first multi-port set of valves for the passage of lysing solution therethrough; e) a precipitating solution container and a conduit which connects said precipitating solution container to an inlet port of said first multi-port set of valves for the passage of precipitating solution therethrough; f) a vacuum pump which is connected to an outlet port of said first multi-port set of valves; g) a bacterial cell suspension container and a conduit which connects said bacterial cell suspension container to an inlet port of said first multi-port set of valves for the passage of bacterial cell suspension therethrough; h) a conduit which connects an outlet port of said first multi-port set of valves to an inlet port of said mixing chamber for the passage of lysing solution, precipitating solution and bacterial cell suspension into said mixing chamber; i) a waste conduit connected to an outlet port of said first multi-port set of valves for the elimination of waste therefrom; j) a second multi-port set of valves having a plurality of inlet and outlet ports for the passage of fluid therethrough; k) a conduit which connects an outlet port of said mixing chamber to an inlet port of said second multi-port set of valves for the passage of fluid therethrough and means for filtering precipitated chromosomal DNA, protein and cellular debris from said fluid which passes therethrough for the production of a filtered fluid which contains dissolved plasmid DNA in the substantial absence of chromosomal DNA, protein and cellular debris; l) a sample collector for collecting the filtered fluid containing the dissolved plasmid DNA; said sample collector having an inlet and an outlet, set inlet of the collector being connected to an outlet port of said second multi-port set of valves for receiving said filtered fluid and said outlet of the colletor being connected to an inlet port of said second multi-port set of valves for returning said filtered fluid to said second multi-port set of valves; m) means for recovering the plasmid DNA, said means for recovering the plasmid DNA being connected by a conduit to an outlet port of said second multi-port set of valves for the passage of filtered fluid from the sample collector to the plasmid DNA recovery means via the second multi-port set of valves; n) a dissolving and cleaning solution container and a conduit which connects said dissolving and cleaning solution container to an inlet port of said second multi-port set of valves for the passage of dissolving and cleaning solution therethrough; o) a conduit which connects the second multi-port set of valves with the mixing chamber for passing the dissolving and cleaning solution from the second multi-port set of valves through said filter means and into the mixing chamber; and p) a conduit which connects an outlet port of said mixing chamber to an inlet port of said first multi-port set of valves for drawing bacterial cell suspension into said mixing chamber by action of said vacuum pump and for the elimination of the dissolving and cleaning solution through the waste conduit via the first multi-port set of valves.