Integrated nucleic acid diagnostic device

a nucleic acid and diagnostic device technology, applied in the field of integrated nucleic acid diagnostic devices, can solve problems such as introducing a potential for error into the overall process

Inactive Publication Date: 2005-11-10
ANDERSON ROLFE C +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0015] The present invention also provides a miniature fluidic system which comporises a differential pressure delivery system for transporting fluids through the system. In particular, in one aspect, the present invention provides a miniature fluidic system, which includes a body having at least a first reaction chamber fluidly connected to a second reaction chamber by a fluid passage. The system also includes a sample inlet, fluidly connected to the first chamber, for introducing a fluid sample into the system. The system further includes a differential pressure delivery system for maintaining the first chamber at a first pressure and the second chamber at a second pressure, wherein the first pressure is greater than ambient pressure and the second pressure is greater than said first pressure. When the second chamber is brought to ambient pressure, the first pressure forces a liquid sample in the first chamber into the second chamber.
[0016] In an alternate aspect, the fluidic system employs a differential pressure delivery source for maintaining the first chamber at a first pressure and the second chamber at a second pressure, where the second pressure is less than ambient pressure and the first pressure is less than the second pressure. When the first chamber is brought to ambient pressure, the second pressure draws a liquid sample in the first chamber into the second chamber.

Problems solved by technology

While current genetic methods are generally capable of identifying these genetic sequences, such methods generally rely on a multiplicity of distinct processes to elucidate the nucleic acid sequences, with each process introducing a potential for error into the overall process.

Method used

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Examples

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example 1

Extraction and Purification of Nucleic Acids

[0198] In separate experiments, HIV cloned DNA was spiked into either horse blood or a suspension of murine plasmacytoma fully differentiated B-cells derived from BALBc mice. Guanidine isothiocyanate was added to a concentration of 4 M, to lyse the material. In separate experiments, the lysate was passed through a cartridge containing glass wool (20 μl), a cartridge with soda glass walls (20 μl), and a glass tube. After 30 minutes at room temperature, the remaining lysate was washed away with several volumes of ethanol:water (1:1) and the captured DNA was eluted at 60° C. using 1×TBE. The yield of eluted DNA was measured using ethidum bromide staining on an agarose gel, and purity was tested by using the eluted material as a template for a PCR reaction. Elution yields ranged from 10% to 25% and PCR yields ranged from 90 to 100% as compared to controls using pure template.

example 2

RNA Preparation Reactions in Miniaturized System

[0199] A model miniature reactor system was designed to investigate the efficacy of miniaturized devices in carrying out prehybridization preparative reactions on target nucleic acids. In particular, a dual reaction chamber system for carrying out in vitro transcription and fragmentation was fabricated. The device employed a tube based structure using a polymer tubing as an in vitro transcription reactor coupled to a glass capillary fragmentation reactor. Reagents not introduced with the sample were provided as dried deposits on the internal surface of the connecting tubing. The experiment was designed to investigate the effects of reaction chamber materials and reaction volume in RNA preparative reaction chambers.

[0200] The sample including the target nucleic acid, DNA amplicons containing a 1 kb portion of the HIV gene flanked with promoter regions for the T3 and T7 RNA primers on the sense and antisense strands, respectively, RNA ...

example 3

PCR Amplification in Miniaturized System

[0203] The miniature polymeric reaction chamber similar to the one described in Example 2 was used for carrying out PCR amplification. In particular, the chamber was fabricated from a planar piece of poycarbonate 4 mm thick, and having a cavity measuring 500 μm deep machined into its surface. A second planar polycarbonate piece was welded over the cavity. This second piece was only 250 μm thick. Thermal control was supplied by applying a peltier heater against the thinner second wall of the cavity.

[0204] Amplification of a target nucleic acid was performed with Perkin-Elmer GeneAmp® PCR kit. The reaction chamber was cycled for 20 seconds at 94° C. (denaturing), 40 seconds at 65° C. (annealing) and 50 seconds at 72° C. (extension). A profile of the thermal cycling is shown in FIG. 9. Amplification of approximately 109 was shown after 35 cycles. FIG. 10C shows production of amplified product in the microchamber as compared to a control using a...

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Abstract

The present invention provides a miniaturized integrated nucleic acid diagnostic device and system. The device of the invention is generally capable of performing one or more sample acquisition and preparation operations, in combination with one or more sample analysis operations. For example, the device can integrate several or all of the operations involved in sample acquisition and storage, sample preparation and sample analysis, within a single integrated unit. The device is useful in a variety of applications, and most notably, nucleic acid based diagnostic applications and de novo sequencing applications.

Description

[0001] This application is a continuation of U.S. application Ser. No. 09 / 751,658, filed on Dec. 31, 2000, now U.S. Pat. 6,830,936, which is a continuation of U.S. patent application Ser. No. 09 / 294,700, filed on Apr. 19, 1999, now U.S. Pat. No. 6,197,595; which is a divisional of U.S. patent application Ser. No. 08 / 671,928, filed on Jun. 27, 1996, now U.S. Pat. No. 5,922,591, which is a continuation-in-part of U.S. patent application Ser. No. 08 / 589,027, filed on Jan. 19, 1996, now U.S. Pat. No. 5,856,174, which claims priority from U.S. Provisional Application 60 / 000,703, filed on Jun. 29, 1995 and also claims priority from U.S. Provisional Application 60 / 000,859, filed on Jul. 3, 1995. application Ser. No. 09 / 751,658 is also a continuation-in-part of U.S. patent application Ser. No. 09 / 210,025, filed on Dec. 11, 1998 now U.S. Pat. No. 6,043,080. The above-identified applications are incorporated herein by reference in their entirety for all purposes.GOVERNMENT RIGHTS [0002] The p...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): B01F11/00C12N15/09B01F11/02B01F13/00B01F13/08B01F15/02B01L3/00B01L7/00C12M1/34C12N1/00G01N35/00G01N35/10
CPCB01F11/0071Y10T436/11B01F13/005B01F13/0059B01F13/08B01F15/0201B01F15/0238B01F2215/0037B01F2215/0073B01L3/5027B01L3/502715B01L3/502723B01L3/50273B01L3/502738B01L3/502746B01L3/502753B01L3/502784B01L7/52B01L9/527B01L2200/0621B01L2200/0684B01L2200/10B01L2300/087B01L2300/0883B01L2300/16B01L2400/0415B01L2400/0421B01L2400/0487B01L2400/049B01L2400/084B01L2400/086G01N35/1095G01N2035/00158Y10T436/25375Y10T436/2575Y10T436/118339Y10T436/25B01F11/0266B01F31/65B01F31/86B01F33/25B01F33/30B01F33/45B01F35/71B01F35/71745B01F2101/23B01F2101/44
Inventor ANDERSON, ROLFE C.LIPSHUTZ, ROBERT J.RAVA, RICHARD P.FODOR, STEPHEN P.A.
Owner ANDERSON ROLFE C
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