| CPC B01L 7/52 (2013.01) [B01L 3/50273 (2013.01); C12Q 1/6844 (2013.01); B01L 3/502723 (2013.01); B01L 2200/10 (2013.01); B01L 2300/0816 (2013.01); B01L 2300/0867 (2013.01); B01L 2300/087 (2013.01); B01L 2300/123 (2013.01); B01L 2300/1822 (2013.01); B01L 2400/0478 (2013.01); B01L 2400/0481 (2013.01); B01L 2400/049 (2013.01); B01L 2400/0644 (2013.01); B01L 2400/065 (2013.01); B01L 2400/0655 (2013.01); B01L 2400/0677 (2013.01); B01L 2400/0683 (2013.01)] | 9 Claims |

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1. A method of nucleic acid extraction and multiplex PCR in a container, comprising:
(a) providing the container having, in fluid communication:
i. one or more ports, including an injector port for introducing a sample into the container, wherein the one or more ports are sealable ports that provide the only access for introduction of a sample into the container,
ii. a cell lysis zone configured for lysing cells or spores located in the sample,
iii. a nucleic acid preparation zone, the nucleic acid preparation zone configured for purifying a plurality of nucleic acids that may be in the sample, wherein the nucleic acid preparation zone, cell lysis zone, and the injector port are fluidically connected; and
iv. at least one amplification zone fluidically connected to the nucleic acid preparation zone, the amplification zone configured for amplification of the plurality of nucleic acids that may be in the sample;
(b) introducing the sample into the container via the injector port and then closing the one or more ports, such that when the one or more ports are closed the container is closed;
(c) lysing cells or spores in the cell lysis zone using motor-driven rotating blades or paddles for bead-milling;
(d) preparing the plurality of nucleic acids that may be in the sample in the nucleic acid preparation zone subsequent to step (c);
(e) moving the plurality of nucleic acids that may be in the sample into the amplification zone to be amplified by a plurality of primer pairs;
(f) thermal cycling the plurality of nucleic acids that may be in the sample in the amplification zone in the presence of PCR reaction components and the plurality of primer pairs to create an amplification mixture; and
(g) detecting which of the plurality of nucleic acids are present in the amplification mixture in the amplification zone.
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