Appeal No. 1995-2839
Application 08/062,023
canceled. Claims 15, 17 and 25 are representative of the subject matter on appeal and
read as follows:
15. A method for the simultaneous amplification and detection of a first target DNA
and a second target DNA comprising:
A) simultaneously subjecting the denatured opposing strands of a first target DNA
and the denatured opposing strands of a second target DNA to polymerase chain reaction
in the presence of:
i) an aqueous composition buffered to a pH of from 7 to 9, and comprising,
in the same solution:
first and second primers which are specific to and hybridizable with,
respectively, first and second nucleic acid sequences which are in opposing strands of a
first target DNA and which are separated from each other along said opposing strands by
from 90 to 400 nucleotides,
third and fourth primers which are specific to and hybridizable with,
respectively, third and fourth nucleic acid sequence which are in opposing strands of a
second target DNA which is the same as or different from said first target DNA, said third
and fourth nucleic acid sequences being different from said first and second nucleic acid
sequences and being separated from each other along said opposing strands of said
second target DNA by from 90 to 400 nucleotides,
each of said first, second, third and fourth primers having a T within
m
the range of from 65 to 74E C, all of said primer T 's being within about 5E C of each other,
m
said first and second primers having nucleotide lengths which differ from each other by no
more than 5 nucleotides, and said third and fourth primers having nucleotide lengths which
differ from each other by no more than 5 nucleotides, and
ii) the additional PCR reagents: a thermostable DNA polymers, a DNA
polymerase cofactor and dNTP's, any or all of said additional PCR reagents being
supplied in the same or a different composition as defined in i),
to simultaneously amplify said opposing first target DNA strands and said opposing
second target DNA strands,
2
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