Multiplex amplification of polynucleotides
AssigneeApplied Biosystems, LLC
InventorsMark R. Andersen, David W. Ruff
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Abstract
The present invention provides methods, reagents and kits for carrying out a variety of assays suitable for analyzing polynucleotides or samples that include an amplification step performed in a multiplex fashion. Also provided are methods for analyzing and improving the efficiency of amplification and for carrying out gene expression analysis.
Claims43
A method for quantifying the expression of target gene sequences of interest in a sample, comprising the steps of:
- (i) amplifying one or more cDNA molecules derived from a sample by polymerase chain reaction in the presence of a plurality of amplification primer sets suitable for amplifying target gene sequence of interest, and in the presence of at least one oligonucleotide probe complementary to a region of an amplified target gene sequence, said at least one oligonucleotide probe optionally labeled with a labeling system suitable for monitoring the amplification reaction as a function of time, and (ii) quantifying the target gene sequences amplified in step (i).
A method for determining a gene expression profile in a sample, comprising the steps of:
- (i) amplifying one or more cDNA molecules derived from said sample by polymerase chain reaction in the presence of a plurality of amplification primer sets suitable for amplifying target gene sequences of interest;
- (ii) identifying amplified target gene sequences having an observed efficiency of amplification greater than a selected level;
- and
- (iii) quantifying the target gene sequences identified in step (ii) to obtain a gene expression profile.
A method of generating a plurality of target sequences of interest, comprising the step of:
- amplifying by polymerase chain reaction one or more target polynucleotides in the presence of a plurality of amplification primers suitable for amplifying target sequences of interest and in the presence of at least one oligonucleotide probe complementary to a region of an amplified target sequence of interest, said oligonucleotide probe being optionally labeled with a labeling system suitable for monitoring an amplification reaction as a function of time.
A method of generating a plurality of different target sequences of interest, comprising the step of:
- amplifying by polymerase chain reaction one or more target polynucleotides in the presence of a plurality of amplification primers suitable for amplifying target sequences of interest and in the presence of a molecule that produces a detectable signal when bound to a double-stranded polynucleotide, said molecule suitable for monitoring the amplification reaction as a function of time, thereby generating a plurality of target sequences of interest.
Dependencies are inferred from the claim text.