Method and means for annealing complementary nucleic acid molecules at an accelerated rate
First Claim
1. The method comprising the steps of binding first nucleic acid molecules to membrane means comprising at least a liquid permeable membrane impermeable to nucleic acid molecules;
- contacting said membrane means and first molecules with an aqueous medium containing second nucleic acid molecules; and
then passing a first electric current through said aqueous medium and said membrane means in a direction to move the second nucleic acid molecules electrophoretically toward said membrane means and into contact with said bound first nucleic acid molecules.
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Accused Products
Abstract
Method and means for accelerating the rate of hybridization of nucleic acid probes with complemental target sequences in probe assays of the filter binding or sandwich filter binding formats. The nucleic acid probes are electrophoretically concentrated at membrane means including at least a dialysis type membrane and to which the target sequences are bound. The rate of hybridization can be further enhanced by means for moving concentrated unannealed probe molecules successively in various directions along the surface of the membrane means to which the target sequences are bound. The invention further includes method and means for electrophoretically removing from the membrane means unannealed probe molecules which became adsorbed to the membrane means during hybridization. In one embodiment of the invention the membrane means comprises a cellulose dialysis membrane in laminate relation with a nitrocellulose or nylon filter having target sequence molecules bound to the surface thereof facing the dialysis membrane. In another embodiment of the invention the membrane means comprises a dialysis membrane to one side of which the target sequence molecules are bound.
178 Citations
38 Claims
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1. The method comprising the steps of binding first nucleic acid molecules to membrane means comprising at least a liquid permeable membrane impermeable to nucleic acid molecules;
- contacting said membrane means and first molecules with an aqueous medium containing second nucleic acid molecules; and
then passing a first electric current through said aqueous medium and said membrane means in a direction to move the second nucleic acid molecules electrophoretically toward said membrane means and into contact with said bound first nucleic acid molecules. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16)
- contacting said membrane means and first molecules with an aqueous medium containing second nucleic acid molecules; and
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17. In combination, a receptacle having partition means separating first and second chambers therein, said partition means comprising membrane means including at least a liquid permeable membrane impervious to nucleic acid molecules;
- electrode means in each of said chambers adapted for connection to a direct current source;
first nucleic acid molecules bound to said membrane means in said first chamber;
a liquid hybridization medium in said first chamber in contact with the electrode means therein, with said membrane means, and with said first molecules;
second nucleic acid molecules in said medium complementary to said first nucleic acid molecules; and
an aqueous second liquid in said second chamber in contact with the electrode means therein and with said membrane means. - View Dependent Claims (18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37)
- electrode means in each of said chambers adapted for connection to a direct current source;
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38. In combination, a receptacle having partition means separating first and second chambers therein, said partition means comprising membrane means including at least a liquid permeable membrane impervious to nucleic acid molecules;
- electrode means in each of said chambers adapted for connection to a direct current source;
first nucleic acid molecules bound to said membrane means in said first chamber; and
aqueous first liquid medium in said first chamber in contact with the electrode means therein, with said membrane means, and with said first molecules;
second nucleic acid molecules in said first liquid medium complementary to said first nucleic acid molecules; and
an aqueous second liquid medium in said second chamber in contact with the electrode means therein and with said membrane means.
- electrode means in each of said chambers adapted for connection to a direct current source;
Specification