# US 2020/0399720 A1 · Compositions and methods for detecting hev nucleic acid

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- Number: US 2020/0399720 A1
- Document ID: US20200399720A1
- Status: Published application
- Kind: A1
- Application number: 17/012,946
- Filed: 2020-09-04
- Published: 2020-12-24
- Earliest claimed: 2013-08-14
- Assignees: Gen-Probe Incorporated
- Applicants: Gen-Probe Incorporated
- Inventors: Kui GAO; Edgar O. ONG; Jennifer COLE; Jeffrey M. Linnen
- Web page: https://patentgrep.com/patent/US20200399720A1

## CPC

- C12Q 1/707: {non-A, non-B Hepatitis, excluding hepatitis D}
- Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

Disclosed are nucleic acid oligomers, including amplification oligomers, capture probes, and detection probes, for detection of Hepatitis E Virus (HEV) nucleic acid. Also disclosed are methods of specific nucleic acid amplification and detection using the disclosed oligomers, as well as corresponding reaction mixtures and kits.

## Claims

### Claim 1

A combination of at least two oligomers for determining the presence or absence of hepatitis E virus (HEV) in a sample, said oligomer combination comprising:
at least two amplification oligomers for amplifying a target region of an HEV target nucleic acid, wherein
(a) at least one amplification oligomer is selected from the group consisting of
(i) an oligomer comprising a target-hybridizing sequence that is from about 14 to about 23 contiguous nucleotides contained in the sequence of SEQ ID NO:63 and that includes at least the sequence of SEQ ID NO:26, including RNA equivalents and DNA/RNA chimerics thereof; and
(ii) an oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:28, including RNA equivalents and DNA/RNA chimerics thereof; and
(b) at least one amplification oligomer is a promoter primer comprising a target-hybridizing sequence consisting of SEQ ID NO:24, SEQ ID NO:45, SEQ ID NO:51, or SEQ ID NO:56, including RNA equivalents and DNA/RNA chimerics thereof and further comprising a promoter sequence joined to the 5′ end of the target-hybridizing sequence of the promoter primer.

### Claim 2 (depends on 1)

The combination of at least two oligomers of claim 1, wherein the at least one amplification oligomer of (a) comprises a target-hybridizing sequence consisting of SEQ ID NO:33, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:61, SEQ ID NO:62, SEQ ID NO:64, SEQ ID NO:65, or SEQ ID NO:66, including RNA equivalents and DNA/RNA chimerics thereof.

### Claim 3 (depends on 1)

The combination of at least two oligomers of claim 1, wherein the at least one amplification oligomer of (a) comprises a target-hybridizing sequence consisting of SEQ ID NO:29, SEQ ID NO:30, or SEQ ID NO:31, including RNA equivalents and DNA/RNA chimerics thereof.

### Claim 4 (depends on 1)

The combination of at least two oligomers of claim 1, wherein said combination comprises a first amplification oligomer as in (b) and a second amplification oligomer as in (b).

### Claim 5 (depends on 1)

The combination of at least two oligomers of claim 1, wherein said combination comprises an amplification oligomer as in (a)(i) and an amplification oligomer as in (a)(ii).

### Claim 6 (depends on 1)

The combination of at least two oligomers of claim 1, wherein the promoter sequence is a T7 promoter sequence.

### Claim 7 (depends on 1)

The combination of at least two oligomers of claim 1, further comprising at least one detectably labeled detection probe oligomer comprising a target-hybridizing sequence that is from about 14 to about 28 nucleotides in length and is configured to specifically hybridize to a target sequence contained within SEQ ID NO:39 or the complement thereof.

### Claim 8 (depends on 7)

The combination of at least two oligomers of claim 7, wherein the detection probe target-hybridizing sequence consists of SEQ ID NO:55 or SEQ ID NO:67, including complements, DNA equivalents, and DNA/RNA chimerics thereof.

### Claim 9 (depends on 7)

The combination of at least two oligomers of claim 7, wherein the at least one detection probe oligomer contains a 2′-methoxy backbone at one or more linkages in the nucleic acid backbone.

### Claim 10

A kit comprising a combination of at least two oligomers for determining the presence or absence of hepatitis E virus (HEV) in a sample, said oligomer combination comprising:
at least two amplification oligomers for amplifying a target region of an HEV target nucleic acid, wherein
(a) at least one amplification oligomer is selected from the group consisting of
(i) an oligomer comprising a target-hybridizing sequence that is from about 14 to about 23 contiguous nucleotides contained in the sequence of SEQ ID NO:63 and that includes at least the sequence of SEQ ID NO:26, including RNA equivalents and DNA/RNA chimerics thereof; and
(ii) an oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:28, including RNA equivalents and DNA/RNA chimerics thereof; and
(b) at least one amplification oligomer is a promoter primer comprising a target-hybridizing sequence consisting of SEQ ID NO:24, SEQ ID NO:45, SEQ ID NO:51, or SEQ ID NO:56, including RNA equivalents and DNA/RNA chimerics thereof and further comprising a promoter sequence joined to the 5′ end of the target-hybridizing sequence of the promoter primer.

### Claim 11

A method for determining the presence or absence of hepatitis E virus (HEV) in a sample, said method comprising:
(1) contacting a sample, said sample suspected of containing HEV, with at least two oligomers for amplifying a target region of an HEV target nucleic acid, said oligomer combination comprising
(a) at least one amplification oligomer comprising a target-hybridizing sequence that is from about 14 to about 23 contiguous nucleotides contained in the sequence of SEQ ID NO:63 and that includes at least the sequence of SEQ ID NO:26, including RNA equivalents and DNA/RNA chimerics thereof; and
(b) at least one amplification oligomer that is a promoter primer comprising a target-hybridizing sequence consisting of SEQ ID NO:24, SEQ ID NO:45, SEQ ID NO:51, or SEQ ID NO:56, including RNA equivalents and DNA/RNA chimerics thereof and further comprising a promoter sequence joined to the 5′ end of the target-hybridizing sequence of the promoter primer;
(2) performing an in vitro nucleic acid amplification reaction, wherein any HEV target nucleic acid present in the sample is used as a template for generating an amplification product; and
(3) contacting the amplification reaction with at least one detectably labeled detection probe oligomer comprising a target-hybridizing sequence that is from about 14 to about 28 nucleotides in length and is configured to specifically hybridize to a target sequence contained within SEQ ID NO:39 or the complement thereof, wherein said contacting is performed under conditions whereby the presence or absence of the amplification product is determined, thereby determining the presence or absence of HEV in the sample.

### Claim 12 (depends on 11)

The method of claim 11, wherein the at least one amplification oligomer of (1)(a) comprises a target-hybridizing sequence consisting of SEQ ID NO:33, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:61, SEQ ID NO:62, SEQ ID NO:64, SEQ ID NO:65, or SEQ ID NO:66, including RNA equivalents and DNA/RNA chimerics thereof.

### Claim 13 (depends on 11)

The method of claim 11, wherein the at least one amplification oligomer of (1)(a) comprises a target-hybridizing sequence consisting of SEQ ID NO:29, SEQ ID NO:30, or SEQ ID NO:31, including RNA equivalents and DNA/RNA chimerics thereof.

### Claim 14 (depends on 11)

The method of claim 11, wherein the promoter sequence is a T7 promoter sequence.

### Claim 15 (depends on 14)

The method of claim 14, wherein the T7 promoter sequence has the sequence shown in SEQ ID NO:73.

### Claim 16 (depends on 11)

The method of claim 11, further comprising purifying the HEV target nucleic acid from other components in the sample before step (1), wherein the purifying step comprises contacting the sample with at least one capture probe oligomer comprising a target-hybridizing sequence covalently attached to a sequence or moiety that binds to an immobilized probe, wherein said target-hybridizing sequence consists of SEQ ID NO:4 or SEQ ID NO:42, including complements, DNA equivalents, and DNA/RNA chimerics thereof.

### Claim 17 (depends on 11)

The method of claim 11, wherein the detection probe target-hybridizing sequence consists of SEQ ID NO:55 or SEQ ID NO:67, including complements, DNA equivalents, and DNA/RNA chimerics thereof.

### Claim 18 (depends on 11)

The method of claim 11, wherein the at least one detection probe oligomer comprises a label selected from the group consisting of
(a) a chemiluminescent label;
(b) a fluorescent label;
(c) a quencher; and
(d) a combination of two or more of (a), (b), and (c).

### Claim 19 (depends on 18)

The method of claim 18, wherein the detecting step (3) detects hybridization of the at least one labeled detection probe oligomer to the amplification product in a homogeneous detection system.

### Claim 20 (depends on 19)

The method of claim 19, wherein the label is a chemiluminescent acridinium ester (AE) compound linked between two nucleobases of the at least one detection probe oligomer.