Method and kit for purifying nucleic acids
Methods for automated extraction of nucleic acids are disclosed. Also disclosed are method and kits for isolating fetal nucleic acids from a plasma sample of a pregnant woman. The method includes flowing the plasma sample through a first filter under conditions that allow binding of the fetal and maternal nucleic acids to the first filter; eluting the fetal and maternal nucleic acids bound to the first filter to produce a concentrated nucleic acid sample; flowing the concentrated nucleic acid sample through a second filter under conditions that allow preferential binding of the maternal nucleic acids to the second filter; and recovering the fetal nucleic acid from the concentrated nucleic acid sample that flow through the second filter.
1. A method for separating and isolating fetal nucleic acids from maternal nucleic acids in a plasma sample, comprising:
(a) flowing a plasma sample comprising fetal nucleic acids and maternal nucleic acids through a first filter, under conditions that allow specific binding of the fetal and maternal nucleic acids to the first filter;
(b) eluting bound fetal and maternal nucleic acids from the first filter to form a concentrated nucleic acid sample comprising fetal nucleic acids and maternal nucleic acids;
(c) flowing the concentrated nucleic acid sample through a second filter, under conditions that allow the maternal nucleic acids to bind to the second filter and the fetal nucleic acids to flow through the second filter; and
(d) collecting the flow-through fraction from the second filter, wherein the flow-through fraction from the second filter contains fetal nucleic acids,
wherein the conditions that allow specific binding of the fetal and maternal nucleic acids to the first filter in step (a) comprise forming a first binding mixture that comprises the plasma sample, an aliphatic alcohol in a range between about 17-25% (v/v) and a chaotropic salt in a concentration range between about 0.5 M to about 4.0 M.
2. The method of claim 1 , wherein the conditions for binding the maternal nucleic acids to the second filter in step (c) comprise forming a second binding mixture that comprises the concentrated nucleic acid sample, an aliphatic alcohol in a range between about 0-10% (v/v) and a chaotropic salt in a concentration range between about 1 M to about 4.0 M.
3. The method of claim 1 , further comprising the steps of:
(e2) flowing the flow-through fraction from the second filter through a third filter under conditions that allow binding of the fetal nucleic acids to the third filter; and
(f2) eluting bound fetal nucleic acids from the third filter.
4. The method of claim 1 , wherein one or both of the first and second filter comprises a glass fit comprising a first section having a first pore size and second section having a second pore size, wherein the first pore size is different from the second pore size.
5. The method of claim 1 , further comprising the steps of:
(e1) eluting bound maternal nucleic acids from the second filter to produce a regenerated second filter;
(f1) flowing the flow-through fraction from the second filter through the regenerated second filter under conditions that allow binding of fetal nucleic acids to the second filter; and
(h1) eluting bound fetal nucleic acids from the second filter in step (f1).
6. The method of claim 5 , wherein the conditions for binding the fetal nucleic acids to the second filter in step (f1) comprise forming a third binding mixture that comprises the flow-through fraction from the second glass frit filter, an aliphatic alcohol in a range between about 10-25% (v/v) and a chaotropic salt in a concentration range between about 1 M to about 5.0 M.
7. The method of claim 1 , wherein the first and second filters are self-supporting glass fits.
8. The method of claim 7 , wherein the glass fits are sintered glass frits.
9. The method of claim 7 , wherein the first glass frit filter has a pore size of 16-40 micron and the second glass frit filter has a pore size of 4-5.5 micron.