Solid-phase nucleic acid isolation
The present invention provides a method of isolating nucleic acid from a sample of cells, said method comprising: (a) binding cells in said sample in a solid support to isolate cells from the sample; (b) lysing the isolated cells; and (c) binding nucleic acid released from said lysed cells to said same solid support and a kit for carrying out such a method. The method may advantageously be used to prepare nucleic acid for use in a nucleic acid-based target cell detection method.
1. A method of isolating nucleic acid from a sample of eukaryotic or prokaryotic cells, said method comprising:
(a) providing a solid support and mixing it with the cell sample and allowing binding of cells in said sample to the solid support to isolate cells from the sample,
(b) separating the cells bound to the solid support from the remainder of the sample by pipetting or decanting, thereby concentrating the cells,
(c) lysing the isolated cells, and
(d) binding nucleic acid released from said lysed cells to a solid support that consists of the same solid support as in step (a); wherein said method further comprises the step of:
(e) eluting nucleic acid from said same solid support; or
(f) detecting the nucleic acid bound to the same solid support.
2. The method of claim 1 , wherein said nucleic acid is DNA.
3. The method of claim 1 , wherein in step (a) the cells are precipitated onto the support using a precipitant.
4. The method of claim 3 , wherein the precipitant comprises alcohol and salt.
5. The method of claim 1 , wherein in step (a) the cells bind to the support by virtue of cell-binding moieties provided on or in the support.
6. The method of claim 5 , wherein said cell-binding moieties permit selective binding of target cells.
7. The method of claim 1 , wherein the support comprises magnetic beads.
8. The method of claim 1 , wherein in step (c), the cells are lysed using a detergent and a chaotrope.
9. The method of claim 1 , wherein in step (c), the cells are lysed using a detergent or a chaotrope.
10. The method of claim 1 , wherein in step (d) the nucleic acid is bound non-specifically to the support.
11. The method of claim 10 , wherein the nucleic acid is precipitated onto the support using a precipitant.
12. The method of claim 11 , wherein the precipitant comprises at least one of alcohol, salt and a detergent.
13. The method of claim 1 , wherein in step (d) the nucleic acid binds to the support by virtue of binding partners provided on the support to assist in the selective capture of nucleic acids.
14. The method of claim 1 , wherein the solid support is in the form of particles, and wherein in step (a) said sample is mixed with said particulate support, and allowed to stand for a suitable interval of time to allow the cells to bind to the support, following which the support is removed from the remainder of the sample.
15. A method for detecting the presence or absence of a target eukaryotic or prokaryotic cell in a sample, said method comprising:
(a) providing a solid support and mixing it with the sample and allowing binding of cells in said sample to the solid support to isolate cells from the sample;
(b) separating the cells bound to the solid support from the remainder of the sample by pipetting or decanting, thereby concentrating the cells,
(c) lysing the isolated cells;
(d) binding nucleic acid released from said lysed cells to a solid support that consists of the same solid support as in step (a); and
(e) detecting the presence or absence of nucleic acid characteristics of said target cells within said bound nucleic acid.