Systems and methods for isolating nucleic acids from cellular samples

• US 10,227,582 B2
• Filed: 05/09/2016
• Issued: 03/12/2019
• Est. Priority Date: 12/22/2011
• Status: Active Grant

• Current Assignee: Ibis Biosciences Incorporated
• Original Assignee: Ibis Biosciences Incorporated
• Inventors: Chiesl, Thomas N., Toro, Michelle, Haupt, Steven Gerald
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US15/150,014
• Publication Number:
  US 20160326513A1
• Time in Patent Office: 1,037 Days
• Field of Search: 536 254, 536 2541, 435 61, 435 911
• CPC Class Codes: C12N 1/06 Lysis of microorganisms
• CPC Class Codes: C12N 1/066 by physical methods
• CPC Class Codes: C12N 15/10 Processes for the isolation, preparation or purification of DNA or RNA chemical preparation of DNA or RNA C07H21/00; preparation of non-structural polynucleotides from microorganisms or with enzymes C12P19/34
• CPC Class Codes: C12N 15/1003 Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
• CPC Class Codes: C12N 15/1006 by means of a solid support carrier, e.g. particles, polymers
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/689 for bacteria
• CPC Class Codes: C12Q 1/6895 for plants, fungi or algae

Abstract

The present disclosure relates to systems and methods for nucleic acid isolation from cellular samples. In particular, the present disclosure provides systems and methods for lysing cells and recovering nucleic acids.

Claims

1. A method of lysing cells, comprising:

a) contacting a biological sample wherein said sample is an aqueous biological fluid comprising a cell with a solution comprising beads and an aromatic solvent wherein said solvent is toluene; and

b) lysing said cells using a bead-beating device in the absence of a buffer.

2. The method of claim 1, wherein said aqueous biological fluid is selected from the group consisting of blood, blood products, and urine.

3. The method of claim 1, wherein said solution further comprises a detergent and or a proteinase.

4. The method of claim 3, wherein said detergent is sodium dodecyl sulfate (SDS).

5. The method of claim 3, wherein said proteinase is proteinase K (PK).

6. The method of claim 1, wherein said solvent is added at a concentration of 50 μ

l to 1 ml per 1 ml of sample.

7. The method of claim 1, further comprising the step of isolating nucleic acids from said sample following said lysing.

8. The method of claim 7, further comprising the step of analyzing said nucleic acids for ample for the presence of a target DNA and/or RNA.

9. The method of claim 8, wherein said target nucleic acid is a pathogen.

10. The method of claim 9, wherein said pathogen is selected from the group consisting of a virus, a fungi, and a bacterium.

11. The method of claim 8, wherein said analyzing comprises a method selected from the group consisting of an amplification method, a sequencing methods, and a hybridization method.

12. A method of lysing cells, comprising:

a) contacting a biological sample wherein said sample is an aqueous biological fluid comprising a cell with a solution comprising beads, a proteinase and an aromatic solvent wherein said solvent is selected from the group consisting of ortho-xylene, meta-xylene and para-xylene; and

b) lysing said cells using a bead-beating device.

13. A method of lysing cells, comprising:

a) contacting a biological sample wherein said sample is an aqueous biological fluid comprising a cell with a solution comprising beads, a proteinase, a detergent and an aromatic solvent; and

b) lysing said cells using a bead-beating device.

14. A method of lysing cells, comprising:

a) contacting a biological sample wherein said sample is an aqueous biological fluid comprising a cell with a solution comprising beads and an aromatic solvent wherein said solvent is selected from the group consisting of ortho-xylene, meta-xylene and para-xylene; and

b) lysing said cells using a bead-beating device in the absence of a buffer.