Method for amplifying quinolone-resistance-determining-regions and identifying polymorphic variants thereof
First Claim
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1. A method for amplifying a polynucleotide sequence of a QRDR comprising the steps of:
- (a) providing a composition comprising a degenerate forward primer of the invention and degenerate reverse primer of the invention, and a sample suspected to have a polynucleotide comprising a QRDR; and
(b) amplifying a QRDR.
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Abstract
The present invention provides a novel method for obtaining regions of bacterial polynucleotide sequence associated with quinolone resistance. In particular, methods for detecting Quinolone Resistance-Determining Regions (QRDRs) across a broad phylogenetic range in prokaryotes are disclosed.
3 Citations
16 Claims
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1. A method for amplifying a polynucleotide sequence of a QRDR comprising the steps of:
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(a) providing a composition comprising a degenerate forward primer of the invention and degenerate reverse primer of the invention, and a sample suspected to have a polynucleotide comprising a QRDR; and
(b) amplifying a QRDR. - View Dependent Claims (2, 3, 4, 5, 11, 12, 13)
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6. A method for identifying a polymorphic polynucleotide sequence of a QRDR comprising the steps of:
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(a) providing a composition comprising a degenerate forward primer of the invention and/or and a degenerate reverse primer of the invention, and a sample suspected to have a polynucleotide comprising a QRDR region;
(b) amplifying a QRDR to obtain an amplified product;
(c) sequencing said amplified product to obtain a first polynucleotide sequence; and
(d) comparing said first polynucleotide sequence with a second polynucleotide sequence of an amplified product made using said degenerate forward primer primer of the invention and said degenerate reverse primer primer of the invention to identify sequence differences between said first polynucleotide sequence and said second polynucleotide sequence. - View Dependent Claims (7, 8, 9, 10)
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- 14. A polynucleotide selected from the group consisting of:
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16. A method for identifying a polynucleotide sequence of a QRDR comprising the steps of:
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(a) providing a composition comprising a degenerate primer of the present invention suitable for use in hybridizations, which comprises a solid surface on which is immobilized at pre-defined regions thereon a plurality of defined oligonucleotide/polynucleotide sequences for hybridization; and
(b) identifying, sequencing, and characterizing genes which are implicated in disease, infection, or development and the use of such identified genes and the proteins encoded thereby in diagnosis, prognosis, therapy and drug discovery.
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Specification