Method of nucleic acid amplification
AssigneeIllumina, Inc.
InventorsEric H. Kawashima, Laurent Farinelli, Pascal Mayer
Life of this patent
Estimating the term…
Abstract
A nucleic acid molecule can be annealed to an appropriate immobilized primer. The primer can then be extended and the molecule and the primer can be separated from one another. The extended primer can then be annealed to another immobilized primer and the other primer can be extended. Both extended primers can then be separated from one another and can be used to provide further extended primers. The process can be repeated to provide amplified, immobilized nucleic acid molecules. These can be used for many different purposes, including sequencing, screening, diagnosis, in situ nucleic acid synthesis, monitoring gene expression, nucleic acid fingerprinting, etc.
Claims25
(canceled)
A multiplex method for determining the presence of one or more target polynucleotide sequences across a plurality of samples, comprising:
- amplifying and tagging target polynucleotides by PCR in each of said plurality of samples with an amplification primer comprising a high throughput sequencing adaptor, a sample-specific tag sequence, and a priming sequence to amplify said target polynucleotide sequence(s);
- combining the amplified polynucleotides and sequencing the polynucleotide pool in high throughput, so as to determine the sequence of at least 100 tagged polynucleotides for each of said samples;
- assigning the nucleotide sequences to the originating samples by the nucleotide sequence of the sample-specific tag, thereby determining the presence of the target polynucleotide sequence(s) across the samples.
A method for simultaneously determining the sequence of one or more sequences of interest in a plurality of samples, comprising:
- amplifying and tagging polynucleotide sequences of interest by PCR in each of said plurality of samples with an amplification primer comprising a sequence complementary to a sequencing primer for high throughput sequencing, a tag sequence which is different for each sample, and primer sequences surrounding the specific sequences of interest to amplify said specific sequences of interest;
- sequencing the amplified polynucleotide sequences of interest from the plurality of samples in high throughput, so as to determine the sequence of over 100 different sequences from the plurality of samples;
- and
- identifying the origin of the amplified polynucleotide sequences of interest using the tag sequence which is different for each sample.
Dependencies are inferred from the claim text.