Electronically mediated nucleic acid amplification in NASBA
First Claim
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1. A method of amplifying specific target nucleic acids in a mixed sample of nucleic acids comprising:
- a. introducing said target nucleic acids onto a bioelectronic microchip having a plurality of electronically addressable capture sites;
b. electronically addressing said target nucleic acids to specific capture sites, said capture sites further having connected thereto capture probes that are specific for said target nucleic acids;
c. hybridizing said target nucleic acids and said amplicons to said capture probes; and
d. subjecting said target nucleic acids to a nucleic acid sequence-based amplification reaction to form amplicons of said target.
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Abstract
A method of improving amplification of nucleic acids using a nucleic acid sequence-based amplification (“NASBA”) method is provided wherein target nucleic acids and NASBA primers are electronically addressed to electronically addressable capture sites of a microchip. This improvement uses electronically induced hybridization of the target nucleic acids to the primers. The primers may be solution-based or immobilized on the capture sites of the microchip.
85 Citations
26 Claims
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1. A method of amplifying specific target nucleic acids in a mixed sample of nucleic acids comprising:
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a. introducing said target nucleic acids onto a bioelectronic microchip having a plurality of electronically addressable capture sites;
b. electronically addressing said target nucleic acids to specific capture sites, said capture sites further having connected thereto capture probes that are specific for said target nucleic acids;
c. hybridizing said target nucleic acids and said amplicons to said capture probes; and
d. subjecting said target nucleic acids to a nucleic acid sequence-based amplification reaction to form amplicons of said target. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8)
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9. A method of amplifying specific target nucleic acids in a mixed sample of nucleic acids comprising:
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a. introducing said target nucleic acids onto a bioelectronic microchip having a plurality of electronically addressable capture sites;
b. electronically addressing said target nucleic acids to specific capture sites, said capture sites further having connected thereto capture probes that are specific for said target nucleic acids;
c. subjecting said target nucleic acids to a nucleic acid sequence-based amplification reaction in solution above said capture sites to form amplicons of said target; and
d. electronically hybridizing said target nucleic acids and said amplicons to said capture probes. - View Dependent Claims (10, 11, 12, 13, 14, 15)
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16. A method for the amplification, multiplex assaying, and detection of target nucleic acids of interest in a mixed sample using a bioelectronic microchip comprising:
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a. introducing at least one of said target nucleic acids of interest onto a bioelectronic microchip having a plurality of electronically addressable capture sites;
b. electronically addressed to said target nucleic acids to any of said plurality of capture sites;
c. amplifying said target nucleic acids to form amplicons of said target nucleic acids;
d. electronically addressing said amplicons to any of said plurality of electronically addressable capture sites;
e. capturing said amplicons and target nucleic acids onto the capture sites to which said amplicons and target nucleic acids are addressed by capture probes that have specificity for binding to said amplicons; and
f. detecting the presence of said captured amplicons and said targets. - View Dependent Claims (17, 18, 19, 20, 21, 22, 23, 24)
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25. A kit for carrying out NASBA-based SDA reactions for use on a bioelectronic microchip comprising:
a. one or more oligonucleotides specific for Factor V, Hemochromotosis, or a bacterium, which oligonucleotides comprise amplification primers, bumper primers, capture probes, and/or signal probes selected from the group consisting of Seq. Id. Nos. 1-62. - View Dependent Claims (26)
Specification