Primers and probes for detection and discrimination of ebola virus

Patent metadata

Field	Value
Publication number	US-2016145698-A1
Application number	US-201514943894-A
Country	US
Kind code	A1
Filing date	Nov 17, 2015
Priority date	Nov 21, 2014
Publication date	May 26, 2016
Grant date	—

Abstract

This invention relates to primers and probes for detecting Ebola virus and one or more subtypes of Ebola virus as well as kits including the probes and primers and methods of using the probes and primers.

First claim

What is claimed is:

1 . A method of determining the presence of a viral hemorrhagic fever virus nucleic acid in a sample comprising:
a) contacting the sample with at least one probe selected from a group consisting of SEQ ID:21, SEQ ID NO:22, SEQ ID NO: 23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO: 27, SEQ ID NO:28, SEQ ID NO:29, or SEQ ID:30 capable of hybridizing under stringent conditions to the viral hemorrhagic fever nucleic acid; and detecting hybridization between the viral hemorrhagic fever nucleic acid and the probe, wherein the detection of hybridization indicates the presence of the viral hemorrhagic fever virus nucleic acid in the sample.

2 . The method of claim 1 , wherein the viral hemorrhagic fever nucleic acid is an Ebola virus nucleic acid.

3 . The method of claim 2 , wherein the Ebola virus nucleic acid is SEQ ID NO:37.

4 . The method of claim 1 , wherein the probe is labeled.

5 . The method of claim 4 , wherein the probe is radiolabeled, or fluorescently labeled.

6 . The method of claim 4 , wherein the probe is labeled with FAM and fluorescent quencher.

7 . The method of claim 1 , further comprising amplifying the viral hemorrhagic fever nucleic acid by polymerase chain reaction (PCR), reverse transcriptase polymerase PCR (RT-PCR), or real time reverse transcriptase PCR. (rt RT-PCR).

8 . The method of claim 7 , wherein the viral hemorrhagic fever nucleic acid is amplified by rt RT-PCR.

9 . The method of claim 8 , wherein the amplifying employs primers that consist of the nucleotide sequences set forth as SEQ ID NO:01, SEQ ID NO:02, SEQ ID NO:03, SEQ ID NO:04, SEQ ID NO:05, SEQ ID NO:06, SEQ ID NO:07, SEQ ID NO:08, SEQ ID NO:09, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, or SEQ ID NO:20.

10 . A primer for the amplification of a viral hemorrhagic fever nucleic acid sequence, wherein the primer consists of a nucleic acid sequence set forth as SEQ ID NO:01, SEQ ID NO:02, SEQ ID NO:03, SEQ ID NO:04, SEQ ID NO:05, SEQ ID NO:06, SEQ ID NO:07, SEQ ID NO:08, SEQ ID NO:09, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, or SEQ ID NO:20.

11 . A set of primers for the amplification of a viral hemorrhagic fever nucleic acid sequence comprising at least one primer according to claim 10 .

12 . The set of primers according to claim 11 , wherein the set of primers comprises:
a. one or more forward primers consisting of the nucleic acid sequence set forth as SEQ ID NO:01, SEQ ID NO:02, SEQ ID NO:03, SEQ ID NO:04, SEQ ID NO:05, SEQ ID NO:06, SEQ ID NO:07, SEQ ID NO:08, SEQ ID NO:09, SEQ ID NO:10 b. one or more reverse primers consisting of the nucleic acid set forth as SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, or SEQ ID NO:20, wherein the set of primers is capable of hybridizing to and directing the amplification of the viral hemorrhagic fever nucleic acid.

13 . The set of primers according to claim 12 , wherein the forward and the reverse primers are a pair of primers, wherein the pair of primers is specific for the amplification of Ebola Zaire virus nucleic acid and consists of the nucleic acid sequence set forth in SEQ ID NO: 37.

14 . A kit for detecting an Ebola nucleic acid comprising:
a) a first oligonucleotide selected from a group consisting of SEQ ID NO:01, SEQ ID NO:02, SEQ ID NO:03, SEQ ID NO:04, SEQ ID NO:05, SEQ ID NO:06, SEQ ID NO:07, SEQ ID NO:08, SEQ ID NO:09, SEQ ID NO:10; a second oligonucleotide selected from a group consisting of SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, or SEQ ID NO:20; a probe selected from the group consisting of SEQ ID:21, SEQ ID NO:22, SEQ ID NO: 23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO: 27, SEQ ID NO:28, SEQ ID NO:29, or SEQ ID:30; a reverse transcriptase; and a polymerase.

Assignees

• Integrated Dna Tech Inc

Inventors

• Rose Scott
• Beltz Kristin

Classifications

• C07K14/005

  from viruses · CPC title

• C12Q1/6827

  for detection of mutation or polymorphism · CPC title

• C12Q1/6883

  for diseases caused by alterations of genetic material · CPC title

• C12Q1/701 Primary

  Specific hybridization probes · CPC title