Amplification using modified primers

• US 6,794,142 B2
• Filed: 10/24/2001
• Issued: 09/21/2004
• Est. Priority Date: 10/25/2000
• Status: Expired due to Term

• Current Assignee: Roche Molecular Systems (Roche Holding AG)
• Original Assignee: Roche Molecular Systems (Roche Holding AG)
• Inventors: Laird, Walter J., Niemiec, John T.
• Primary Examiner(s): Horlick, Kenneth R.
• Assistant Examiner(s): Tung, Joyce
• Application Number:
  US10/083,233
• Publication Number:
  US 20030044817A1
• Time in Patent Office: 1,063 Days
• Field of Search: 435/6, 435/91.2, 435/810, 536/24.3, 536/24.33, 536/23.1, 536/25.31, 536/25.32, 536/26.71
• US Class Current: 435/6.1
• CPC Class Codes: C12Q 1/6848 characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
• CPC Class Codes: C12Q 1/6853 using modified primers or templates
• CPC Class Codes: C12Q 2525/117 incorporating modified base

Abstract

The present invention provides modified primers for use in the amplification of a nucleic acid sequence. Amplifications carried out using the modified primers result in less template-independent non-specific product (primer dimer) compared to amplifications carried out using unmodified primers.

Claims

1. A kit for carrying out a nucleic acid amplification reaction, wherein said kit comprises a pair of primers, wherein a least one primer of said pair contains a modified nucleotide within the three 3′

terminal nucleotide positions;

wherein said modified nucleotide is selected from the group consisting of 2′

-O-methyl nucleotides, 2′

-fluoro-nucleotides, 2′

-amino nucleotides, and arabinose nucleotides.

2. The kit of claim 1, wherein said modified nucleotide is a 2′

-O-methyl nucleotide.

3. The kit of claim 1, wherein said modified nucleotide is a 2′

-fluoro-nucleotide.

4. The kit of claim 1, wherein said modified nucleotide is a 2′

-amino nucleotide.

5. The kit of claim 1, wherein said modified nucleotide is an arabinose nucleotide.

6. The kit of claim 2, wherein said modified nucleotide is at the 3′

terminal position.

7. The kit of claim 3, wherein said modified nucleotide is at the 3′

terminal position.

8. The kit of claim 4, wherein said modified nucleotide is at the 3′

terminal position.

9. The kit of claim 5, wherein said modified nucleotide is at the 3′

terminal position.

10. A kit of claim 1, wherein each primer of said pair of primers independently contains a modified nucleotide within the three 3′

terminal nucleotide positions;

wherein said modified nucleotide is selected from the group consisting of 2′

-O-methyl nucleotides, 2′

-fluoro-nucleotides, 2′

-amino nucleotides, and arabinose nucleotides.

11. A method for amplifying a nucleic acid target sequence, wherein said method comprises carrying out a primer-based amplification reaction in a reaction mixture comprising a pair of primers, wherein a least one primer of said pair contains a modified nucleotide within the three 3′

terminal nucleotide positions;

wherein said modified nucleotide is selected from the group consisting of 2′

-O-methyl nucleotides, 2′

-fluoro-nucleotides, 2′

-amino nucleotides, and arabinose nucleotides.

12. The method of claim 11, wherein said modified nucleotide is a 2′

-O-methyl nucleotide.

13. The method of claim 11, wherein said modified nucleotide is a 2′

-fluoro-nucleotide.

14. The method of claim 11, wherein said modified nucleotide is a 2′

-amino nucleotide.

15. The method of claim 11, wherein said modified nucleotide is an arabinose nucleotide.

16. The method of claim 12, wherein said modified nucleotide is at the 3′

terminal position.

17. The method of claim 13, wherein said modified nucleotide is at the 3′

terminal position.

18. The method of claim 14, wherein said modified nucleotide is at the 3′

terminal position.

19. The method of claim 15, wherein said modified nucleotide is at the 3′

terminal position.

20. A method of claim 11, wherein each primer of said pair of primers independently contains a modified nucleotide within the three 3′

terminal nucleotide positions;

wherein said modified nucleotide is selected from the group consisting of 2′

-O-methyl nucleotides, 2′

-fluoro-nucleotides, 2′

-amino nucleotides, and arabinose nucleotides.