Amplification of nucleic acids with electronic detection

DETAILED DESCRIPTION In accordance with the objects outlined above, the present invention provides methods for detecting a target nucleic acid sequence.
The methods comprise hybridizing at least a first primer nucleic acid to the target sequence to form a first hybridization complex, and contacting the first hybridization complex with a first enzyme to form a modified first primer nucleic acid.
The first hybridization complex is then disassociated.
These steps may be repeated a plurality of times.
A first assay complex is then formed comprising at least one ETM and the modified first primer nucleic acid.
The assay complex is covalently attached to an electrode.
Electron transfer is then detected between the ETM and the electrode as an indication of the presence of the target sequence.
The method can include the same method on a second target sequence substantially complementary to the the first target sequence.
In an additional aspect, the method utilizes a DNA polymerase and the modification to the primer is an extension of the primer such that the polymerase chain reaction (PCR) occurs.
In a further aspect, the method utilizes a ligase and the modification to the primer comprises a ligation of the first primer which hybridizes to a first domain of the first target sequence to a third primer which hybridizes to a second adjacent domain of the first target sequence, such that the ligase chain reaction (LCR) occurs.
In an additional aspect, the method utilizes a first primer comprising a first probe sequence, a first scissile linkage and a second probe sequence.
The enzyme will cleave the first scissile linkage resulting in the separation of the first and the second probe sequences and the disassociation of the hybridization complex while leaving the first target sequence intact, such that the cycling probe technology (CPT) reaction occurs.
In a further aspect, the method utilizes a first enzyme that is a polymerase that extends the first primer to form a modified first primer comprising a first newly synthesized strand, and said method further comprises the addition of a second enzyme comprising a nicking enzyme that nicks the extended first primer leaving the first target sequence intact