CN118166163A - Multiple PCR detection kit for high-risk human papilloma virus - Google Patents
Multiple PCR detection kit for high-risk human papilloma virus Download PDFInfo
- Publication number
- CN118166163A CN118166163A CN202410453247.2A CN202410453247A CN118166163A CN 118166163 A CN118166163 A CN 118166163A CN 202410453247 A CN202410453247 A CN 202410453247A CN 118166163 A CN118166163 A CN 118166163A
- Authority
- CN
- China
- Prior art keywords
- hpv type
- type
- hpv
- detecting
- seq
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000001514 detection method Methods 0.000 title claims abstract description 34
- 241000701806 Human papillomavirus Species 0.000 title claims abstract description 19
- 238000006243 chemical reaction Methods 0.000 claims abstract description 18
- 238000007403 mPCR Methods 0.000 claims abstract description 14
- 238000011144 upstream manufacturing Methods 0.000 claims description 20
- 239000000203 mixture Substances 0.000 claims description 17
- 238000000034 method Methods 0.000 claims description 11
- 108020004707 nucleic acids Proteins 0.000 claims description 11
- 150000007523 nucleic acids Chemical class 0.000 claims description 11
- 102000039446 nucleic acids Human genes 0.000 claims description 11
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 claims description 10
- 238000012163 sequencing technique Methods 0.000 claims description 9
- 230000003321 amplification Effects 0.000 claims description 6
- 238000003199 nucleic acid amplification method Methods 0.000 claims description 6
- 238000012165 high-throughput sequencing Methods 0.000 claims description 5
- 229910052697 platinum Inorganic materials 0.000 claims description 5
- 241000341655 Human papillomavirus type 16 Species 0.000 claims description 4
- 238000002360 preparation method Methods 0.000 claims description 3
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 claims description 2
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 claims description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 2
- 208000022361 Human papillomavirus infectious disease Diseases 0.000 abstract description 42
- 230000008696 hypoxemic pulmonary vasoconstriction Effects 0.000 abstract description 42
- 108020004414 DNA Proteins 0.000 description 7
- 238000012408 PCR amplification Methods 0.000 description 5
- 238000010276 construction Methods 0.000 description 4
- 238000012070 whole genome sequencing analysis Methods 0.000 description 4
- 206010008342 Cervix carcinoma Diseases 0.000 description 3
- 208000009608 Papillomavirus Infections Diseases 0.000 description 3
- 102000006382 Ribonucleases Human genes 0.000 description 3
- 108010083644 Ribonucleases Proteins 0.000 description 3
- 101150061589 Rpp30 gene Proteins 0.000 description 3
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 description 3
- 201000010881 cervical cancer Diseases 0.000 description 3
- 238000013461 design Methods 0.000 description 3
- 239000012634 fragment Substances 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 206010059313 Anogenital warts Diseases 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 210000000056 organ Anatomy 0.000 description 2
- 206010001197 Adenocarcinoma of the cervix Diseases 0.000 description 1
- 208000034246 Adenocarcinoma of the cervix uteri Diseases 0.000 description 1
- 206010008263 Cervical dysplasia Diseases 0.000 description 1
- 108020004638 Circular DNA Proteins 0.000 description 1
- 208000000907 Condylomata Acuminata Diseases 0.000 description 1
- 241000450599 DNA viruses Species 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 208000000260 Warts Diseases 0.000 description 1
- 208000025009 anogenital human papillomavirus infection Diseases 0.000 description 1
- 201000004201 anogenital venereal wart Diseases 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 201000006662 cervical adenocarcinoma Diseases 0.000 description 1
- 208000007951 cervical intraepithelial neoplasia Diseases 0.000 description 1
- 239000000539 dimer Substances 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 210000002919 epithelial cell Anatomy 0.000 description 1
- 230000009841 epithelial lesion Effects 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- 210000004392 genitalia Anatomy 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 231100000590 oncogenic Toxicity 0.000 description 1
- 230000002246 oncogenic effect Effects 0.000 description 1
- 238000003753 real-time PCR Methods 0.000 description 1
- 230000001850 reproductive effect Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 201000010153 skin papilloma Diseases 0.000 description 1
- 206010041823 squamous cell carcinoma Diseases 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
- C12Q1/701—Specific hybridization probes
- C12Q1/708—Specific hybridization probes for papilloma
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/686—Polymerase chain reaction [PCR]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/112—Disease subtyping, staging or classification
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Immunology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Analytical Chemistry (AREA)
- Physics & Mathematics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Genetics & Genomics (AREA)
- Virology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses a multiplex PCR detection kit for high-risk human papilloma viruses. The detection kit can realize accurate typing by using only 10 pairs of primers on the premise of covering 18 high-risk HPVs (HPV 16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, 73 and 82) on the premise of completing detection of a sample in a single reaction, and is suitable for various NGS platforms. Meanwhile, the result interpretation is completed by the sequencer, so that the result interpretation is not easily influenced by personal subjective factors, and the result is more accurate and objective.
Description
Technical Field
The invention relates to the technical field of molecular biology, in particular to a multiplex PCR detection kit for high-risk human papilloma viruses.
Background
Human papillomaviruses (Human papillomavirus, HPV) are a generic name for a class of double-stranded circular DNA viruses that transduce and parasitize epithelial cells in the human reproductive organ or other tissue organs. HPV infection can cause genital warts or cutaneous warts, and can induce cervical cancer. It has been reported that almost all cervical squamous carcinomas and more than 85% of cervical adenocarcinomas are caused by HPV infection.
The research at present finds that HPV exists in hundreds of different subtypes and has obvious regional distribution characteristics. HPV is generally classified into two major categories, high-risk and low-risk, based on the oncogenic potential of each subtype. Low-risk HPV can often cause condyloma acuminatum or cervical epithelial lesions, rarely causing cervical cancer. High-risk HPV infection can cause male and female external genitalia cancer, high cervical intraepithelial neoplasia, and cervical cancer. Numerous studies have demonstrated that 18 subtypes, 16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, 73 and 82 are the most common high-risk HPV types.
Accordingly, there is a need in the art for comprehensive detection of high-risk HPV types. The detection of HPV by fluorescent quantitative PCR technology is widely used at present. Unfortunately, this technique is often not amenable to accurate typing and is disadvantageous for statistical analysis of the data. In addition, the method needs to set a plurality of reactions for detecting one sample, and has higher requirements on the quality of the sample.
HPV accurate typing can be performed on multiple samples simultaneously using high throughput sequencing (NGS), with lower sample quality requirements. However, the effective sequencing length of NGS is typically around 100bp, such that the currently mature multiplex PCR primer combinations cannot be directly used.
Therefore, developing a multiplex PCR primer combination capable of specifically amplifying 18 high-risk HPVs and directly applied to the technical system of the NGS platform is a problem to be solved in the field.
In view of this, the present invention has been made.
Disclosure of Invention
The invention aims to provide a multiplex PCR detection kit for high-risk human papilloma viruses, which can be used for simultaneously detecting 18 high-risk human papilloma virus subtypes.
The invention is realized in the following way:
In a first aspect, the invention provides a nucleic acid composition for detecting human papillomavirus comprising primers for detecting HPV type 16, HPV type 18, HPV type 26, HPV type 31, HPV type 33, HPV type 35, HPV type 39, HPV type 45, HPV type 51, HPV type 52, HPV type 53, HPV type 56, HPV type 58, HPV type 59, HPV type 66, HPV type 68, HPV type 73 and HPV type 82;
The primer includes: the upstream and downstream primers for detecting HPV16/35 type are shown as SEQ ID NO. 1-2;
the upstream and downstream primers for detecting HPV18/45 type are shown as SEQ ID NO. 3-4;
The upstream and downstream primers for detecting HPV31 type are shown as SEQ ID NO. 5-6;
the upstream and downstream primers for detecting HPV33/58 type are shown as SEQ ID NO. 7-8;
the upstream and downstream primers for detecting HPV39/59/68 type are shown as SEQ ID NO. 9-10;
the upstream and downstream primers for detecting HPV51/82 type are shown as SEQ ID NO. 11-12;
The upstream and downstream primers for detecting HPV52/58 type are shown as SEQ ID NO. 13-14;
the upstream and downstream primers for detecting HPV53/56/66 type are shown as SEQ ID NO. 15-16;
The upstream and downstream primers for detecting HPV26 type are shown as SEQ ID NO. 17-18;
the upstream and downstream primers for detecting HPV73 type are shown as SEQ ID NO. 19-20.
The nucleic acid composition consists of 10 pairs of primers, which can be applied to 18 types of HPV detection, wherein 7 pairs of primers can simultaneously amplify 2 or 3 subtypes. By means of the combining strategy, compared with the primer combination amplified one by one, the nucleic acid composition provided by the invention can reduce the complexity of the primers, avoid mutual interference caused by excessive primers, and reduce the cost to a certain extent.
In some embodiments, dG in the primer is modified to 8-OXO-dG and part dT is replaced with dU.
In the invention, the inventor carries out base modification on the primers, and the modified primer combination can be compatible with NGS library building kits widely used in the market at present so as to carry out high-throughput sequencing on subsequent amplification products.
In a second aspect, the invention also provides application of the nucleic acid composition in preparing a kit for detecting human papillomavirus.
The nucleic acid composition of the invention is mainly applied to 18 HPV subtypes which are most common at present: in the detection of HPV16, HPV18, HPV26, HPV31, HPV33, HPV35, HPV39, HPV45, HPV51, HPV52, HPV53, HPV56, HPV58, HPV59, HPV66, HPV68, HPV73 and HPV 82.
In a third aspect, the invention also provides a kit for detecting human papillomavirus, comprising the nucleic acid composition.
In some embodiments, the final concentration of each primer in the detection system is 0.1. Mu. Mol/L to 0.5. Mu. Mol/L.
In some embodiments, the detection system further comprises an internal reference primer, the sequence of which is shown in SEQ ID NOS.21-22, the internal reference gene being the ribonuclease RPP30 gene.
In some embodiments, the detection system further comprises a2× Platinum Multiplex PCR MASTER Mix containing DNA polymerase, dNTPs, mg 2+, PCR buffer.
In a fourth aspect, the invention also provides a method for detecting human papillomavirus for non-diagnostic purposes, comprising: extracting DNA of a sample to be detected as a PCR reaction template, adding the PCR reaction template into a reaction system to perform multiplex PCR reaction, and then carrying out high-throughput sequencing on an amplification result and analyzing HPV information in the sequencing result to obtain HPV genotype of the sample.
In some embodiments, the reaction system is 25 μl: 2X Platinum Multiplex PCR MASTER Mix 12.5. Mu.L, the final concentration of each of the above primers was 0.25. Mu. Mol/L, 2. Mu.L of template DNA, and the mixture was made up to 25. Mu.L with enzyme-free water.
The invention has the following beneficial effects:
The detection kit can realize accurate typing by using only 10 pairs of primers on the premise of covering 18 high-risk HPVs on the whole surface, and can finish detection of samples in a single reaction. The amplified fragments are below 200bp, subsequent operations such as enzyme digestion and the like are not needed, and the amplified fragments can be directly used as initial templates for NGS library construction and are suitable for various NGS platforms. Meanwhile, the result interpretation is completed by the sequencer, so that the result interpretation is not easily influenced by personal subjective factors, and the result is more accurate and objective.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention more clear, the technical solutions of the embodiments of the present invention will be clearly and completely described below. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
The invention provides a group of detection primers suitable for typing high-risk human papilloma viruses HPV16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, 73 and 82, the detection primers are reasonably designed and preferably obtained, and the detection primers have good specificity, low cost and high flux when used for PCR amplification, can be used for typing a plurality of high-risk viruses at the same time, and have simple and easy operation detection procedures. Meanwhile, the invention optimizes the PCR amplification reaction and further improves the amplification efficiency.
The features and capabilities of the present invention are described in further detail below in connection with the examples.
Example 1
The present example is a multiplex PCR primer design, specifically as follows:
Sequences of 18 high-risk HPVs are obtained from NCBI database, and primer pairs complementarily paired with the DNA sequences of 18 HPVs are designed according to the base complementation pairing principle. When designing primers, the problems of dimer formation between primers, non-specific amplification and the like are also required to be avoided, and meanwhile, the design of the primers becomes particularly critical due to the limitation of the second generation sequencing read length. In consideration of various factors, 10 pairs of primers are designed for 18 high-risk HPVs, and simultaneously, a ribonuclease RPP30 gene is used as an internal reference gene to find out a ribonuclease RPP30 gene conservation section, so that a pair of primer combinations are designed. The information of the primers designed by the invention is shown in Table 1:
TABLE 1 primer sequence information
DG in the primer is modified into 8-OXO-dG; the partial dT is replaced with dU, and the replaced dU is underlined. Y represents C or T; m represents A or C.
The total of 22 PCR primers obtained by the design method is shown in SEQ ID NO. 1-SEQ ID NO. 22, the concentration of each PCR primer is prepared to be 20 mu mol/L, and the primer mixture is prepared by the equal volume ratio.
Example 2
The embodiment provides a kit for detecting human papilloma virus and application, wherein the kit comprises 10 pairs of primers and a pair of reference genes designed for 18 high-risk type HPVs provided in the embodiment 1, and further comprises 2X Platinum Multiplex PCR MASTER Mix.
The application method of the kit comprises the following steps:
(1) Preparation of DNA templates
(2) PCR amplification of DNA templates
First, a PCR reaction system was prepared as shown in Table 2:
TABLE 2 multiplex PCR System
PCR amplification was then performed, and the reaction procedure for amplification is shown in Table 3:
TABLE 3 multiplex PCR procedure
(3) NGS detection
Library construction was performed using VAHTS AMPSEQ Library Prep Kit and analysis was performed based on the illuminea Hiseq2000 sequencer. The analysis parameter is single-end reading length of 75bp.
And comparing each sequencing result with the sequence of each HPV subtype in NCBI to obtain a detection result.
Example 3
The present example is an application of the kit provided in example 2, namely, the kit of example 2 is used to detect 15 clinical samples, and specifically the following steps are included:
(1) Preparation of DNA templates
GeneRotex 96 full-automatic nucleic acid instrument (medical instrument record: shanxi mechanical equipment 20150054) and matched reagent based on Samsung technology production extract DNA of sample, and the operation flow is carried out according to the instruction provided by the manufacturer.
(2) PCR amplification of DNA templates
The reaction system is shown in Table 2, and the reaction procedure is shown in Table 3.
(3) NGS detection
1) Library construction
Library construction was performed using VAHTS AMPSEQ Library Prep Kit V kit (cat# NA 210-01) produced by Northenan Biotechnology, and experimental procedures were performed according to the instructions provided by the manufacturer.
2) High throughput sequencing
Sequencing is carried out based on an illuminea Hiseq2000 high-throughput sequencer, a single-ended sequencing mode is selected, the fragment read length is 75bp, and the total sequencing throughput is 130M. The experimental procedure was performed according to the standard procedure provided by the manufacturer.
(4) Sequencing results
The 15 samples in this example were analyzed based on whole genome sequencing, and the whole genome sequencing results were compared with the detection results of this example, and the results are shown in table 4:
TABLE 4 detection results
As shown in Table 4, the results of the test in this example showed good agreement with whole genome sequencing. Wherein samples 1-6, 8-12, 14 and 15 are identical to the whole genome sequencing results, samples 7 and 13 do not have both subtypes in the results after detection, since the HPV subtypes referred to in the present invention do not comprise HPV11 and HPV41 subtypes. The detection primer and the detection method can be embodied from the detection results, have good detection effect, and can specifically identify 18 HPV subtypes.
The above description is only of the preferred embodiments of the present invention and is not intended to limit the present invention, but various modifications and variations can be made to the present invention by those skilled in the art. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (10)
1. A nucleic acid composition for detecting human papillomavirus comprising primers for detecting HPV type 16, HPV type 18, HPV type 26, HPV type 31, HPV type 33, HPV type 35, HPV type 39, HPV type 45, HPV type 51, HPV type 52, HPV type 53, HPV type 56, HPV type 58, HPV type 59, HPV type 66, HPV type 68, HPV type 73 and HPV type 82;
The primer comprises: the upstream and downstream primers for detecting HPV16/35 type are shown as SEQ ID NO. 1-2;
the upstream and downstream primers for detecting HPV18/45 type are shown as SEQ ID NO. 3-4;
The upstream and downstream primers for detecting HPV31 type are shown as SEQ ID NO. 5-6;
the upstream and downstream primers for detecting HPV33/58 type are shown as SEQ ID NO. 7-8;
the upstream and downstream primers for detecting HPV39/59/68 type are shown as SEQ ID NO. 9-10;
the upstream and downstream primers for detecting HPV51/82 type are shown as SEQ ID NO. 11-12;
The upstream and downstream primers for detecting HPV52/58 type are shown as SEQ ID NO. 13-14;
the upstream and downstream primers for detecting HPV53/56/66 type are shown as SEQ ID NO. 15-16;
The upstream and downstream primers for detecting HPV26 type are shown as SEQ ID NO. 17-18;
the upstream and downstream primers for detecting HPV73 type are shown as SEQ ID NO. 19-20.
2. The nucleic acid composition of claim 1, wherein dG in the primer is modified to 8-OXO-dG and part dT is replaced with dU.
3. Use of the nucleic acid composition of claim 1 or 2 for the preparation of a kit for detecting human papillomavirus.
4. A use according to claim 3, wherein the use comprises detection of HPV type 16, HPV type 18, HPV type 26, HPV type 31, HPV type 33, HPV type 35, HPV type 39, HPV type 45, HPV type 51, HPV type 52, HPV type 53, HPV type 56, HPV type 58, HPV type 59, HPV type 66, HPV type 68, HPV type 73 and HPV type 82 of human papillomavirus.
5. A kit for detecting human papillomavirus comprising the nucleic acid composition of claim 1 or 2.
6. The kit according to claim 5, wherein the final concentration of each primer in the detection system is 0.1. Mu. Mol/L to 0.5. Mu. Mol/L.
7. The kit of claim 6, wherein the detection system further comprises an internal reference primer having a sequence as set forth in SEQ ID NOS.21-22.
8. The kit of claim 7, wherein the detection system further comprises a2 x Platinum Multiplex PCR MASTER Mix containing DNA polymerase, dNTPs, mg 2+, PCR buffer.
9. A method for detecting human papillomavirus for non-diagnostic purposes, comprising: and taking the sample extracted DNA as a PCR reaction template, adding the PCR reaction template into a reaction system to perform multiple PCR reactions, and then carrying out high-throughput sequencing on the amplification result and analyzing HPV information in the sequencing result to obtain the HPV genotype of the sample.
10. The method of claim 9, wherein the reaction system is 25 μl: 2X Platinum Multiplex PCR MASTER Mix 12.5. Mu.L, the final concentration of each primer according to claim 1 or 2 is 0.25. Mu. Mol/L, 2. Mu.L of template DNA, and the primers are made up to 25. Mu.L with enzyme-free water.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410453247.2A CN118166163A (en) | 2024-04-16 | 2024-04-16 | Multiple PCR detection kit for high-risk human papilloma virus |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410453247.2A CN118166163A (en) | 2024-04-16 | 2024-04-16 | Multiple PCR detection kit for high-risk human papilloma virus |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN118166163A true CN118166163A (en) | 2024-06-11 |
Family
ID=91350505
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410453247.2A Pending CN118166163A (en) | 2024-04-16 | 2024-04-16 | Multiple PCR detection kit for high-risk human papilloma virus |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN118166163A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN118773384A (en) * | 2024-09-09 | 2024-10-15 | 广州凯普医学检验所有限公司 | A human papillomavirus typing primer set for high-throughput sequencing of targeted pathogens and its screening method |
-
2024
- 2024-04-16 CN CN202410453247.2A patent/CN118166163A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN118773384A (en) * | 2024-09-09 | 2024-10-15 | 广州凯普医学检验所有限公司 | A human papillomavirus typing primer set for high-throughput sequencing of targeted pathogens and its screening method |
| CN118773384B (en) * | 2024-09-09 | 2025-01-07 | 广州凯普医学检验所有限公司 | Typing primer group of human papilloma virus for targeting pathogen high-throughput sequencing and screening method thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Poljak et al. | Hybrid Capture II HPV Test detects at least 15 human papillomavirus genotypes not included in its current high-risk probe cocktail | |
| Zaravinos et al. | Molecular detection methods of human papillomavirus (HPV) | |
| CN101435002B (en) | A method for detecting human papillomavirus genotype | |
| CN105603121A (en) | Method, oligonucleotide and kit for detecting high-risk HPV | |
| CN115612744A (en) | Human papillomavirus typing and related gene methylation integrated detection model and its construction method | |
| CN118360437B (en) | Early screening and monitoring kit for HPV (human papilloma Virus) related tumors | |
| CN116769964A (en) | Primer, primer probe composition and kit for detecting high-risk human papilloma virus | |
| JP2008524990A (en) | Detection of human papillomavirus | |
| CN114164305B (en) | A primer and typing probe combination for detecting human papillomavirus and its application | |
| CN116121465A (en) | Application of primer and its composition with probe in preparation of test kit for detecting HPV | |
| CN114875181A (en) | Quality control product for human papilloma virus molecular detection and preparation method thereof | |
| CN101886137B (en) | Fluorescent quantitative PCR kit for qualitative detection of high-risk HPV | |
| CN118166163A (en) | Multiple PCR detection kit for high-risk human papilloma virus | |
| CN102367490A (en) | Method for detecting viruses | |
| CN106048081A (en) | HPV (human papilloma virus) typing detection primers as well as detection method and application thereof | |
| CN101942440A (en) | Primers and method for detecting and typing human papilloma viruses in esophagi | |
| CN111172328A (en) | Complete set of primers, complete set of probes, kit and method for HPV nucleic acid typing detection | |
| CN108179226B (en) | Nucleic acid composition for detecting human papilloma virus, application thereof and kit | |
| Dictor et al. | Single-tube multiplex PCR using type-specific E6/E7 primers and capillary electrophoresis genotypes 21 human papillomaviruses in neoplasia | |
| KR101761701B1 (en) | HPV Specific Probe and DNA Chip for Detecting Genetic Type of HPV Containing Thereof | |
| CN116042920A (en) | NGS detection method and kit for tiny residual focus of cervical cancer patient after treatment based on targeting HPV | |
| WO2008074182A1 (en) | Method for the detection and typing of human papillomavirus, and the reagent kit thereof | |
| CN108728578A (en) | Detect the method and kit of a variety of high-risk human mammilla papillomavirus and its hypotype simultaneously in single tube reaction | |
| CN117327734A (en) | Positive quality control product for genotyping detection of high-risk human papilloma virus, and preparation method and application thereof | |
| Ju et al. | Asymmetric GP5+/6+ PCR and hybridization with fluorescence polarization assay of 15 human papillomavirus genotypes in clinical samples |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |