Method and kit for dna isolation
AssigneeGlobal Life Sciences Solutions Germany GmbH
InventorsRichard Martin West, Angela Sian Davies, Malcolm John Hatcher, Monika Seidel
Life of this patent
Estimating the term…
Abstract
A method for isolating cell-free DNA from liquid body sample, comprising the following steps: a) Providing liquid body sample; b) Adding to said sample: a solid phase capable of binding DNA; a binding buffer comprising a detergent and a chaotropic agent; and 2-propanol, to form a binding mixture thereof; c) Washing the solid phase to remove unbound material; and d) Eluting bound cell-free DNA, wherein the majority of the eluted DNA is <400 bp.
Claims26
A method for isolating cell-free DNA from liquid body sample, comprising the following steps:
- a) Providing liquid body sample;
- b) Adding to said sample:
- a solid phase capable of binding DNA;
- a binding buffer comprising a detergent and a chaotropic agent;
- and
- 2-propanol,
- to form a binding mixture thereof;
- c) Washing the solid phase to remove unbound material;
- and
- d) Eluting bound cell-free DNA,
- wherein the majority of the eluted DNA is <400 bp.
A method for size-selective isolation of cell-free DNA from liquid body sample, comprising the following steps:
- a) Providing liquid body sample;
- b) Adding to said sample:
- an aqueous suspension of silica coated magnetic microbeads capable of binding DNA;
- a binding buffer comprising guanidinium thiocyanate and non-ionic surfactant such as Triton X-100;
- and
- 2-propanol,
- to form a binding mixture thereof such that said binding mixture comprises the non-ionic surfactant such as Triton X-100 at around 20-30% w/v, guanidinium thiocyanate at around 1.5-2.5 M and 2-propanol at around 15-25% v/v;
- c) Incubating the binding mixture at room temperature for about 10-30 minutes to promote binding of cell-free DNA to the magnetic microbeads;
- d) Washing the magnetic microbeads with one or more wash buffers comprising ethanol;
- e) Adding elution buffer to the washed magnetic beads of step d) to release the cell-free DNA bound to the magnetic microbeads in solution;
- and
- f) Optionally analysing or quantifying the cell-free DNA obtained in step e).
Use of guanidinium thiocyanate and Triton X-100 to form a binding buffer composition for size-selective binding of cell-free DNA present in blood plasma to silica-coated magnetic microbeads formulated in an aqueous suspension at 20-200 mg/ml, wherein said binding buffer is intended to be brought into contact with 2-propanol, blood plasma and magnetic microbeads to form a binding mixture comprising:
- around 1.5-2.5 M guanidinium thiocyanate;
- around 20-30% w/v of Triton X-100;
- around 15-25% v/v of 2-propanol;
- and around 25-40% v/v of blood plasma.
Kit comprising silica coated microbeads capable of binding 50-400 bp DNA from a liquid body sample in the presence of guanidinium thiocyanate, Triton X-100 and 2-propanol.
Dependencies are inferred from the claim text.