Compositions for use in identification of alphaviruses

• US 8,119,336 B2
• Filed: 03/02/2005
• Issued: 02/21/2012
• Est. Priority Date: 03/03/2004
• Status: Expired due to Fees

• Current Assignee: Ibis Biosciences Incorporated
• Original Assignee: Ibis Biosciences Incorporated
• Inventors: Sampath, Rangarajan, Hall, Thomas A., Eshoo, Mark W.
• Primary Examiner(s): MUMMERT, STEPHANIE KANE
• Application Number:
  US11/070,632
• Publication Number:
  US 20100035227A1
• Time in Patent Office: 2,547 Days
• Field of Search: None
• US Class Current: 435/5
• CPC Class Codes: G16B 25/00 ICT specially adapted for hybridisation; ICT specially adapted for gene or protein expression
• CPC Class Codes: G16B 25/20 Polymerase chain reaction [PCR]; Primer or probe design; Probe optimisation
• CPC Class Codes: G16B 30/00 ICT specially adapted for sequence analysis involving nucleotides or amino acids
• CPC Class Codes: G16B 30/10 Sequence alignment; Homology search
• CPC Class Codes: G16B 40/00 ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
• CPC Class Codes: G16B 40/10 Signal processing, e.g. from mass spectrometry [MS] or from PCR

Abstract

The present invention provides oligonucleotide primers and compositions and kits containing the same for rapid identification of alphaviruses by amplification of a segment of viral nucleic acid followed by molecular mass analysis.

Claims

1. A composition comprising a purified oligonucleotide primer pair configured to generate amplicons from two or more members of the alphavirus genus by hybridizing a forward primer and a reverse primer to conserved regions of a nsP1 encoding gene in two or more members of said alphavirus genus, said primer pair comprising a forward primer with 100% sequence identity with SEQ ID NO:

93 and a reverse primer with 100% sequence identity with SEQ ID NO;

66.

2. The composition of claim 1 wherein either or both of said oligonucleotide primers comprises at least one modified nucleobase.

3. The composition of claim 1 wherein either or both of said oligonucleotide primers comprises a non-templated T residue on the 5′

end.

4. The composition of claim 1 wherein either or both of said oligonucleotide primers comprises at least one non-template tag.

5. The composition of claim 1 wherein either or both of said oligonucleotide primers comprises at least one molecular mass modifying tag.

6. A kit comprising the composition of claim 1.

7. The kit of claim 6 further comprising at least one calibration polynucleotide.

8. The kit of claim 6 further comprising at least one ion exchange resin linked to magnetic beads.

10. A method for generating alphavirus amplicons comprising:

1) providing a composition according to claim 1; and

2) contacting an alphavirus nucleic acid with the composition to generate alphavirus amplicons.

9. A composition comprising a purified oligonucleotide primer pair configured to generate amplicons from two or more members of the alphavirus genus by hybridizing a forward primer and a reverse primer to conserved regions of a nsP1 encoding gene in two or more members of said alphavirus genus, said primer pair comprising a forward primer with 100% sequence identity with SEQ ID NO:

21 and a reverse primer with 100% sequence identity with SEQ ID NO;

66.