CN104774974A - Coliphage MS2 standard sample and preparing method thereof - Google Patents
Coliphage MS2 standard sample and preparing method thereof Download PDFInfo
- Publication number
- CN104774974A CN104774974A CN201510227340.2A CN201510227340A CN104774974A CN 104774974 A CN104774974 A CN 104774974A CN 201510227340 A CN201510227340 A CN 201510227340A CN 104774974 A CN104774974 A CN 104774974A
- Authority
- CN
- China
- Prior art keywords
- phage
- sample
- escherichia coli
- standard model
- pfu
- 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
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
-
- 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]
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Immunology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biophysics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Physics & Mathematics (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 coliphage MS2 standard sample. The concentration of the coliphage MS2 standard sample is 1.57*1011 pfu/mL, and the standard sample can be used as a quality control substance for the detection process when food-borne RNA viruses are detected. According to a preparing method of the coliphage MS2 standard sample, evenness and stability are tested, and the guarantee period is 12 months.
Description
Technical field:
The invention belongs to the technical field that bioanalysis test, clinical medical inspection and food source property RNA viruses detect, be specifically related to a kind of Escherichia coli MS 2 phage standard model.
Background technology:
Phage MS2 belongs to the bacteriophage of acellular protista, obligate infects and parasitizes in corresponding bacterial body alive, there is the primary condition as enterovirus indicator in water body: do not have pathogenic to people, and in the water surrounding having enterovirus to pollute ubiquity; High density inoculation and test in place can be carried out; Quantity is higher than enterovirus.Morphological specificity, physicochemical property, in resistance of natural environmental condition and sterilizing agent etc. and enterovirus similar; Detection operation has the advantages such as easy quick, safe and reliable, affected by environment little and equipment is simple.
With the host of other viruses as compared with the mankind, animal and plant etc., phage MS2 has following advantage: (1) under lab, phage MS2 is easy to fast culture, does not specially require the space in laboratory, facilities and equipment, and high density preparation (can reach 10
10~ 10
12pFU/mL).(2) with traditional indicator organism of enterovirus as compared with coliform, streptococcus faecium etc., the biological structure of phage MS2, moiety, copy mode are more similar to enterovirus with sterilization resistance etc.The detection to dissimilar water sample such as B.Benoit shows, educable enterovirus and phage MS2 quantitatively exist good corresponding relation.(3) phage ubiquity in the water body having enterovirus to pollute, and quantity is a lot of, at least suitable with enterovirus with the resistibility of disinfecting to water body purification; It can not breed in water simultaneously, does not have pathogenic, can be detected by simple, quick, cheap method.
Along with developing rapidly of Protocols in Molecular Biology, with polymerase chain reaction technique (po|ymerase chain reaction, PCR) the various diagnostic technique in molecular biology based on, because of its there is high specificity, highly sensitive, linearity range is wide, the advantage such as reproducible, quantitatively accurate, convenient and swift, become the research means of most worthy in biomedical sector, be widely used in the detection of clinical samples.In detection of nucleic acids process, the accuracy of molecular biology for detection is easily subject to the impact of following many factors: (1) instrument temperature control program is incorrect, or can not accurate temperature controlling; (2) reversed transcriptive enzyme and Taq enzyme activity is low, reaction solution is of poor quality, primer, probe design are unreasonable; (3) nucleic acid-templated extracted amount is few, the object segment degradation of detection, residual ethanol and phenol amount excessive, sample Impurity removal is clean, and as the cholate in stool sample, the urea in the protoheme in blood and urine can produce restraining effect to PCR reaction; (4) nucleic acid sequence morphs; (5) misoperation of experiment operator.Therefore, do nucleic acid particularly RNA amplification experiment time, need strict quality control method, namely need specific Quality Control thing, meanwhile, quantitative assay will be carried out to nucleic acid, usually also must have nucleic acid standard substance.Quality Control thing and standard substance must have following characteristics: (1) easily prepares; (2) in storage and use procedure, there is enough stability; (3) lifeless matter infectivity; (4) whole process detected can be monitored, reliable results.Current RNA viruses Quality Control thing mainly contains exposed RNA fragment, virion and armor R (armored RNA).Escherichia coli MS 2 phage is envelope protein package structure RNA structure, with RNA viruses structural similitude, and MS2 no pathogenicity; In food source property RNA viruses detection system, add Escherichia coli MS 2 phage as inner quality control thing, be that RNA viruses detects ideal inner quality control material.Thus the Quality Control thing that can be used as of this biolobic material is to monitor the whole testing processes such as virion lysis efficiency and nucleic acid extraction, compensate for the deficiency of naked rna fragment as Quality Control thing and standard substance.
Summary of the invention:
Technical problem to be solved by this invention is to provide a kind of Escherichia coli MS 2 phage reference material, and concentration is 1.57 × 10
11pfu/mL; This reference material can be used as testing process Quality Control material in food source property RNA viruses testing process.
The present invention solves its technical problem to adopt following technical scheme:
A preparation for Escherichia coli MS 2 phage reference material, comprises the following steps:
(1) preparation of Host Strains
First choose 1 ATCC 15597 strain Escherichia coli with transfering loop and be seeded in the cultivation of TSA-YE plate overnight; Picking 1 transfering loop bacterium in 10mL LB liquid nutrient medium, overnight incubation;
(2) prepared by a large amount of phage MS2
1, the Host Strains nutrient solution () prepared is inoculated in LB liquid nutrient medium according to 0.5% (volume ratio), 36 ± 1 DEG C, after 150rpm shaking culture 4-5h, in the ratio of 0.5-1% (volume ratio) by phage (10
8~ 10
9pfu/mL) be inoculated in above-mentioned inoculum, guarantee that the ratio of phage number and bacterium number is 0.1 ~ 2.0.
2, after inoculating, inoculum is at 36 ± 1 DEG C, and 150rpm shaking culture 5h or spend the night gets nutrient solution centrifugal 10min under 10 000g, collects supernatant liquor; Namely the phage suspensions of purifying is obtained with the membrane filtration that aperture is 0.22 μm.
3, tiring of phage is measured
Adopt double layer agar method.Concrete operations are as follows:
Phage suspensions to be measured is done 10 times of serial dilutions to 10
10doubly, get the bottom nutrient agar medium that Host Strains nutrient solution 0.2mL adds to each plate respectively, draw suitable dilution phage suspensions 0.1mL to add to each plate respectively and mix with Host Strains, pour the top-layer agar that 2 ~ 3mL is chilled to 50 DEG C into, mixing, solidify latter 37 DEG C and be inverted cultivation 18 ~ 24h, each dilution gradient does 3 repetitions, calculates plaque number.The mean value calculation of getting the plate of plaque number between 30 ~ 300 is tired:
To tire (pfu/mL)=Mean plaque number × extension rate × 10
4, the dilution of phage
Adopt suspension medium (SM) to dilute phage solution, and be sub-packed in 2mL cell cryopreservation tube and preserve.
(3) stability test
Different time points is got 25 DEG C, 4 DEG C ,-80 DEG C sample 3 pipes preserved respectively and is carried out phage titer mensuration, and carries out the stability of each preservation condition of statistical study; 25 DEG C, 4 DEG C sample times continue 6 months, and sampling in every 1 month once;-80 DEG C of sample times continue 24 months, and sampling in every 2 months once.
(4) uniformity test
Get-80 DEG C of sample 15 pipes preserved and carry out phage titer mensuration, and carry out statistical study, the homogeneity of evaluation criteria sample.
(5) standard model definite value
Result according to uniformity test carries out uncertainty evaluation, the definite value of column criterion of going forward side by side sample, and this Escherichia coli MS 2 phage standard model concentration is 1.57 × 10
11pfu/mL.
Beneficial effect of the present invention:
Phage MS2 prepared by the present invention can be used as testing process Quality Control material, homogeneity and good stability, height of tiring as in food source property RNA viruses testing process.
Accompanying drawing illustrates:
Fig. 1 is that the embodiment of the present invention 1 Escherichia coli MS 2 phage is tired test pattern.
Fig. 2 is the embodiment of the present invention 2 Escherichia coli MS 2 phage real-time fluorescence RT-PCR typical curve. ordinate zou is Ct value, and X-coordinate is Escherichia coli MS 2 phage concentration (pfu/mL).
Embodiment:
Also the present invention is set forth further by reference to the accompanying drawings below by specific embodiment.
Embodiment 1: Escherichia coli MS 2 phage preparation of standard sample
(1) preparation of Host Strains
First choose 1 ATCC 15597 strain Escherichia coli with transfering loop and be seeded in the cultivation of TSA-YE plate overnight; Picking 1 transfering loop bacterium in 10mL LB liquid nutrient medium, overnight incubation;
(2) prepared by a large amount of phage MS2
1, the Host Strains nutrient solution () prepared is inoculated in LB liquid nutrient medium according to 0.5% (volume ratio), 36 ± 1 DEG C, after 150rpm shaking culture 4-5h, in the ratio of 0.5-1% (volume ratio) by phage (10
8~ 10
9pfu/mL) be inoculated in above-mentioned inoculum, guarantee that the ratio of phage number and bacterium number is 0.1 ~ 2.0.
2, after inoculating, inoculum is at 36 ± 1 DEG C, and 150rpm shaking culture 5h or spend the night gets nutrient solution centrifugal 10min under 10 000g, collects supernatant liquor; Namely the phage suspensions of purifying is obtained with the membrane filtration that aperture is 0.22 μm.
3, tiring of phage is measured
Adopt double layer agar method.Concrete operations are as follows:
Phage suspensions to be measured is done 10 times of serial dilutions to 10
10doubly, get the bottom nutrient agar medium that Host Strains nutrient solution 0.2mL adds to each plate respectively, draw suitable dilution phage suspensions 0.1mL to add to each plate respectively and mix with Host Strains, pour the top-layer agar that 2 ~ 3mL is chilled to 50 DEG C into, mixing, solidify latter 37 DEG C and be inverted cultivation 18 ~ 24h, each dilution gradient does 3 repetitions, calculates plaque number.The mean value calculation of getting the plate of plaque number between 30 ~ 300 is tired:
To tire (pfu/mL)=Mean plaque number × extension rate × 10
4, the dilution of phage
Adopt suspension medium (SM) to dilute phage solution, and be sub-packed in 2mL cell cryopreservation tube and preserve.
(3) uniformity test
Get-80 DEG C of sample 10 pipes preserved and carry out phage titer mensuration, and carry out statistical study, the homogeneity of evaluation criteria sample.
(4) stability test
Different time points is got-20 DEG C of sample 2 pipes preserved respectively and is carried out phage titer mensuration, and carries out statistical study assessment stability, is respectively 2,4,6,8,10,12,14,16,18,20 months sample time.
(5) standard model definite value
Result according to uniformity test carries out uncertainty evaluation, the definite value of column criterion of going forward side by side sample, and this Escherichia coli MS 2 phage standard model concentration is 1.57 × 10
11pfu/mL.
Embodiment 2: the structure of real-time fluorescence PCR typical curve
(1) specimen material: by the standard model (1.57 × 10 of preparation
11pfu/mL).ABI Ambion Life TechnologiesAM1005 AgPath-ID One-Step RT-PCR Kit。ABI 7500 FAST real-time fluorescence quantitative PCR instrument.
(2) structure of real-time fluorescence PCR typical curve
By the standard model (1.57 × 10 of preparation
11pfu/mL).10 times of gradient dilution to 2.5 × 10
9pfu/mL, gets 2.5 × 10
9pfu/mL sample 200 μ L, cracking 5min under 95 DEG C of conditions, then 10 times of gradient dilutions to 10 ×, 100 ×, 1000 ×, 10000 ×, then corresponding respectively phage MS2 concentration is 1.57 × 10
8pfu/mL, 1.57 × 10
7pfu/mL, 1.57 × 10
6pfu/mL, 1.57 × 10
5pfu/mL.
The preparation of RT-PCR premixed liquid is carried out according to the explanation of RT-PCR kit, each reaction system is 25 μ L, and comprise the sample of premixed liquid 20 μ L and 5 μ L, each extent of dilution repeats 3 times, and select on instrument and standard model option is set, according to phage MS2 concentration assignment.RT-PCR reaction parameter is: 45 DEG C, 10min; 95 DEG C, 10min; 50 circulations: 94 DEG C, 15sec; 60 DEG C, 45sec.Instrument generates typical curve (see Fig. 3) automatically according to assignment and Ct value: y=58.716-3.352lnx (y is Ct value, and x is Escherichia coli MS 2 phage concentration (pfu/mL)).
Embodiment 3: take MS2 as the detection that quality control of procedure standard model carries out Norwalk virus
(1) specimen material: buy Ark Shell, chlamys farreri, clam, each 1 part of oyster from Tuan Dao market, Qingdao.
(2) open shellfish shell with tweezers, scalpel etc., separating digesting gland (digestive glands), often kind of shellfish collects 2g ± 0.2 respectively in 50mL centrifuge tube, operates on ice, is shredded by visceral mass by sterile scissors.
(3) Proteinase K working fluid and the 100 μ l process control materials (2.5 × 10 of 2mL are added respectively
8pfu/mL phage MS2 solution), whirlpool mixes.
(4) 37 DEG C of shaking tables hatch 1h, 320r/min.
(5) 60 DEG C of water-bath 15min.
(6) 4 DEG C of centrifugal 5min of 3000g, get supernatant liquor and record cumulative volume, get 500 μ L supernatant liquors and extract RNA, be finally dissolved in the elutriant of 100 μ L, detect in order to PCR.
(7) preparation of RT-PCR premixed liquid is carried out according to the explanation of RT-PCR kit, each reaction system is 25 μ L, comprises the sample of premixed liquid 20 μ L and 5 μ L, and each sample makees the RNA extracting solution of former times and 10 times dilution respectively, RT-PCR reaction parameter is: 45 DEG C, 10min; 95 DEG C, 10min; 50 circulations: 94 DEG C, 15sec; 60 DEG C, 45sec.
(8) according to the Ct value of RT-PCR, calculate the concentration of virus in extracting solution according to typical curve, infer extraction efficiency.
(9) show according to extraction efficiency result: extraction efficiency is between 3.70%-7.84%, and the organic efficiency meeting ISO 15216:2-2013 is greater than the requirement of 1%.
Table 1 Escherichia coli MS 2 phage adds recovery test
| Sample number into spectrum | Sample ID | Extraction efficiency |
| 1 | Ark Shell | 7.84％ |
| 2 | Chlamys farreri | 7.88％ |
| 3 | Clam | 3.78％ |
| 4 | Oyster | 3.70％ |
Embodiment 4: the uniformity test of Escherichia coli MS 2 phage standard model
Get-80 DEG C of sample 10 pipes preserved to carry out carrying out titration according to the phage titer measuring method in embodiment 1 (two), and carry out statistical study, the homogeneity of evaluation criteria sample, result shows (see table 2): F=0.0515 < F threshold value=4.4139, between each pipe, difference is not remarkable, has good uniformity.
Table 2 analysis of Uniformity
Embodiment 5: the stability analysis of Escherichia coli MS 2 phage standard model
Different time points is got-20 DEG C of sample 2 pipes preserved respectively and is carried out phage titer mensuration, averages and carries out statistical study assessment stability, be respectively 2,4,6,8,10,12,14,16,18,20 months sample time.Result shows (see table 3):
Table 3 stability analysis (analysis of variance in regression)
The concentration monitor of table 3-a Escherichia coli MS 2 phage
| -20 DEG C of shelf times (moon) | Record the concentration (pfu/mL) of Escherichia coli MS 2 phage |
| 2 | 1.55E+11 |
| 4 | 1.57E+11 |
| 6 | 1.59E+11 |
| 8 | 1.57E+11 |
| 10 | 1.59E+11 |
| 12 | 1.62E+11 |
| 14 | 1.58E+11 |
| 16 | 1.57E+11 |
| 18 | 1.55E+11 |
| 20 | 1.56E+11 |
The analysis of variance in regression of the concentration of table 3-b Escherichia coli MS 2 phage
| Difference source | Sum of squares (SS) | Degree of freedom (df) | Square root of the variance (MS) | F value | F threshold value |
| Regression analysis | 1 | 0.677871148 | 0.677871148 | 0.0165 | 0.9011 |
| Residual error | 8 | 329.3221289 | 41.16526611 | ||
| Amount to | 9 | 330 |
F=0.0165 < F threshold value=0.9011, show that each sample time MS2 change in concentration difference is not obvious, in 20 months ,-20 DEG C have good stability, therefore its quality guaranteed period is decided to be 12 months is feasible.
Claims (3)
1. an Escherichia coli MS 2 phage standard model, is characterized in that: concentration is 1.57 × 10
11pfu/mL; This standard model can be used as testing process Quality Control material in food source property RNA viruses testing process.
2. a preparation for Escherichia coli MS 2 phage standard model, is characterized in that, comprises the following steps:
(1) preparation of Host Strains
First choose 1 ATCC 15597 strain Escherichia coli with transfering loop and be seeded in the cultivation of TSA-YE plate overnight; Picking 1 transfering loop bacterium in 10mL LB liquid nutrient medium, overnight incubation;
(2) prepared by a large amount of phage MS2
1, the Host Strains nutrient solution () prepared is according to 0.5%(volume ratio) be inoculated in LB liquid nutrient medium, 36 ± 1 DEG C, after 150rpm shaking culture 4-5h, in 0.5-1%(volume ratio) ratio phage is inoculated in above-mentioned inoculum, guarantee that the ratio of phage number and bacterium number is 0.1 ~ 2.0;
2, after inoculating, inoculum is at 36 ± 1 DEG C, and 150rpm shaking culture 5 h or spend the night gets nutrient solution centrifugal 10 min under 10 000 g, collects supernatant liquor; Namely the phage suspensions of purifying is obtained with the membrane filtration that aperture is 0.22 μm;
3, tiring of phage is measured
Adopt double layer agar method, concrete operations are as follows:
Phage suspensions to be measured is done 10 times of serial dilutions to 10
10doubly, get the bottom nutrient agar medium that Host Strains nutrient solution 0.2mL adds to each plate respectively, draw suitable dilution phage suspensions 0.1mL to add to each plate respectively and mix with Host Strains, pour the top-layer agar that 2 ~ 3 mL are chilled to 50 DEG C into, mixing, solidify latter 37 DEG C and be inverted cultivation 18 ~ 24 h, each dilution gradient does 3 repetitions, calculates plaque number; The mean value calculation of getting the plate of plaque number between 30 ~ 300 is tired:
To tire (pfu/mL)=Mean plaque number × extension rate × 10
4, the dilution of phage
Adopt suspension medium (SM) to dilute phage solution, and be sub-packed in 2mL cell cryopreservation tube and preserve;
(3) stability test
Different time points is got 25 DEG C, 4 DEG C ,-80 DEG C sample 3 pipes preserved respectively and is carried out phage titer mensuration, and carries out the stability of each preservation condition of statistical study; 25 DEG C, 4 DEG C sample times continue 6 months, and sampling in every 1 month once;-80 DEG C of sample times continue 24 months, and sampling in every 2 months once;
(4) uniformity test
Get-80 DEG C of sample 15 pipes preserved and carry out phage titer mensuration, and carry out statistical study, the homogeneity of evaluation criteria sample;
(5) standard model definite value
Result according to uniformity test carries out uncertainty evaluation, the definite value of column criterion of going forward side by side sample, and this Escherichia coli MS 2 phage standard model concentration is 1.57 × 10
11pfu/mL.
3. the application of Escherichia coli MS 2 phage standard model in food source property RNA viruses testing process.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510227340.2A CN104774974A (en) | 2015-05-07 | 2015-05-07 | Coliphage MS2 standard sample and preparing method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510227340.2A CN104774974A (en) | 2015-05-07 | 2015-05-07 | Coliphage MS2 standard sample and preparing method thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN104774974A true CN104774974A (en) | 2015-07-15 |
Family
ID=53616738
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510227340.2A Pending CN104774974A (en) | 2015-05-07 | 2015-05-07 | Coliphage MS2 standard sample and preparing method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104774974A (en) |
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105219737A (en) * | 2015-10-29 | 2016-01-06 | 广州呼研所生物技术有限公司 | The preparation of quality control product, application and test kit is marked in Escherichia coli M13 phage |
| CN105238765A (en) * | 2015-10-29 | 2016-01-13 | 广州呼研所生物技术有限公司 | Preparation and application of coliphage MS2 internal standard quality control product and kit |
| CN105462973A (en) * | 2016-01-14 | 2016-04-06 | 山东出入境检验检疫局检验检疫技术中心 | Preparation method for GII Norovirus virus-like particles and application thereof |
| CN105505926A (en) * | 2016-01-14 | 2016-04-20 | 山东出入境检验检疫局检验检疫技术中心 | Preparing method for HAV virus-like particle and application thereof |
| CN106011081A (en) * | 2016-06-14 | 2016-10-12 | 苏州埃瑞特生物技术有限公司 | Large-scale production method of bacteriophage |
| CN106085974A (en) * | 2016-06-07 | 2016-11-09 | 博奥生物集团有限公司 | A kind of zika virus pseudovirion and preparation method thereof |
| CN106085967A (en) * | 2016-06-14 | 2016-11-09 | 苏州埃瑞特生物技术有限公司 | A kind of production method of high-titer phage |
| CN107741502A (en) * | 2017-10-12 | 2018-02-27 | 中国农业科学院农业质量标准与检测技术研究所 | A kind of preparation method of carbon, nitrogen stable isotope standard sample in beef protein |
| CN108676779A (en) * | 2018-04-25 | 2018-10-19 | 广州市微生物研究所 | A method of detection air clearing product purifies air pnagus medius ability |
| CN109097487A (en) * | 2018-03-06 | 2018-12-28 | 广东省微生物研究所（广东省微生物分析检测中心） | It is a kind of wash one's hands or wash product remove virus effectiveness in vitro test method |
| CN110904276A (en) * | 2019-12-23 | 2020-03-24 | 苏州良辰生物医药科技有限公司 | Application of bacteriophage in verification of virus removal process of biological product |
| CN113930544A (en) * | 2018-01-30 | 2022-01-14 | 中国计量科学研究院 | Method for valuing virus standard substance for evaluating protection performance of personnel protection equipment |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103497962A (en) * | 2013-10-15 | 2014-01-08 | 上海市计量测试技术研究院 | Plasmid standard molecular adaptable to sakazakii real-time fluorescent quantitation PCR (polymerase chain reaction) detection |
-
2015
- 2015-05-07 CN CN201510227340.2A patent/CN104774974A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103497962A (en) * | 2013-10-15 | 2014-01-08 | 上海市计量测试技术研究院 | Plasmid standard molecular adaptable to sakazakii real-time fluorescent quantitation PCR (polymerase chain reaction) detection |
Non-Patent Citations (4)
| Title |
|---|
| SHULMAN L M ET AL.: "Evaluation of four different systems for extraction of RNA from stool suspensions using MS-2 coliphage as an exogenous control for RT-PCR inhibition", 《PLOS ONE》 * |
| 任衍菊等: "生物防护服穿透性噬菌体指标筛选", 《中国公共卫生》 * |
| 温和心等: "RNA病毒质控物研究进展", 《动物医学进展》 * |
| 王秋英等: "噬菌体MS2φX174 的双层琼脂平板和液体培养基扩增方法的建立", 《土壤》 * |
Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105219737A (en) * | 2015-10-29 | 2016-01-06 | 广州呼研所生物技术有限公司 | The preparation of quality control product, application and test kit is marked in Escherichia coli M13 phage |
| CN105238765A (en) * | 2015-10-29 | 2016-01-13 | 广州呼研所生物技术有限公司 | Preparation and application of coliphage MS2 internal standard quality control product and kit |
| CN105462973A (en) * | 2016-01-14 | 2016-04-06 | 山东出入境检验检疫局检验检疫技术中心 | Preparation method for GII Norovirus virus-like particles and application thereof |
| CN105505926A (en) * | 2016-01-14 | 2016-04-20 | 山东出入境检验检疫局检验检疫技术中心 | Preparing method for HAV virus-like particle and application thereof |
| CN106085974B (en) * | 2016-06-07 | 2019-08-09 | 博奥生物集团有限公司 | A kind of Zika virus pseudovirion particle and preparation method thereof |
| CN106085974A (en) * | 2016-06-07 | 2016-11-09 | 博奥生物集团有限公司 | A kind of zika virus pseudovirion and preparation method thereof |
| CN106085967A (en) * | 2016-06-14 | 2016-11-09 | 苏州埃瑞特生物技术有限公司 | A kind of production method of high-titer phage |
| CN106011081A (en) * | 2016-06-14 | 2016-10-12 | 苏州埃瑞特生物技术有限公司 | Large-scale production method of bacteriophage |
| CN107741502A (en) * | 2017-10-12 | 2018-02-27 | 中国农业科学院农业质量标准与检测技术研究所 | A kind of preparation method of carbon, nitrogen stable isotope standard sample in beef protein |
| CN113930544A (en) * | 2018-01-30 | 2022-01-14 | 中国计量科学研究院 | Method for valuing virus standard substance for evaluating protection performance of personnel protection equipment |
| CN113930544B (en) * | 2018-01-30 | 2024-07-05 | 中国计量科学研究院 | Method for determining virus standard substance for evaluating protective performance of personnel protective equipment |
| CN109097487A (en) * | 2018-03-06 | 2018-12-28 | 广东省微生物研究所（广东省微生物分析检测中心） | It is a kind of wash one's hands or wash product remove virus effectiveness in vitro test method |
| CN109097487B (en) * | 2018-03-06 | 2021-09-07 | 广东省微生物研究所（广东省微生物分析检测中心） | An in-vitro test method for the virus-removing effect of hand-washing or hand-washing products |
| CN108676779A (en) * | 2018-04-25 | 2018-10-19 | 广州市微生物研究所 | A method of detection air clearing product purifies air pnagus medius ability |
| CN110904276A (en) * | 2019-12-23 | 2020-03-24 | 苏州良辰生物医药科技有限公司 | Application of bacteriophage in verification of virus removal process of biological product |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104774974A (en) | Coliphage MS2 standard sample and preparing method thereof | |
| Birch et al. | A comparison of nucleic acid amplification techniques for the assessment of bacterial viability | |
| RU2520082C2 (en) | Method of bacterial ghost (bg) | |
| CN101144775B (en) | Bacterial real-time fluorescent quantitative polymerase chain reaction detection kit | |
| Tomaso et al. | Comparison of commercial DNA preparation kits for the detection of Brucellae in tissue using quantitative real-time PCR | |
| McNerney et al. | Mycobacteriophage and their application to disease control | |
| CN103421896A (en) | RNA isothermal amplification nucleic acid detection kit aiming at Escherichia coli 0157 | |
| Zhang et al. | Diagnostics and detection of African swine fever virus | |
| CN104152582B (en) | Ox lumpy skin disease virus Taqman-MGB probe for real-time fluorescence is primer, kit and detection method for quantitative PCR detection | |
| Holmdahl et al. | Hydrogen peroxide vapour treatment inactivates norovirus but has limited effect on post-treatment viral RNA levels | |
| CN105238765A (en) | Preparation and application of coliphage MS2 internal standard quality control product and kit | |
| CN101372713B (en) | Method for detecting Francisella tularensis using multiple PCR technology | |
| KR101009321B1 (en) | How to quickly detect infectious microorganisms in unknown samples | |
| US11098380B2 (en) | Reagent and method for rapid detection of porcine adenovirus | |
| CN106435007A (en) | Primer, probe, kit and method for detecting bacillus erysipelatos-suis by fluorescent quantitative PCR (Polymerase Chain Reaction) | |
| CN103602727B (en) | Detection primer set, detection kit and detection method for rapidly detecting Ralstonia solanacearum | |
| Mozioğlu et al. | Oligomer based real-time detection of microorganisms producing nuclease enzymes | |
| RU2730658C1 (en) | Kit for detecting pathogenic microorganisms of listeria monocytogenes type and method for detection thereof in samples of biomaterial, in samples of fodders, in objects of external environment | |
| CN103571942A (en) | Isothermal amplification method of vibrio parahaemolyticus VP nucleic acid | |
| CN107746894B (en) | Method for rapidly identifying nucleotide point mutation of phytophthora sojae β -tubulin gene and resistance of phytophthora sojae β -tubulin gene to ethaboxam | |
| CN110184371B (en) | A kind of specific primer for detecting Acinetobacter pietii and its method and application | |
| JP7248329B2 (en) | Method for judging the risk of developing onion vegetative disease | |
| JP2020068678A (en) | How to detect Streptococcus mutans | |
| CN104894232A (en) | Citrobacter freundii fluorescent quantitative PCR diagnostic reagent kit | |
| CN101705298A (en) | Quick bacterium examination kit and detection method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| EXSB | Decision made by sipo to initiate substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20150715 |