CN105695556A - Screw inoculation method - Google Patents
Screw inoculation method Download PDFInfo
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- CN105695556A CN105695556A CN201610090903.2A CN201610090903A CN105695556A CN 105695556 A CN105695556 A CN 105695556A CN 201610090903 A CN201610090903 A CN 201610090903A CN 105695556 A CN105695556 A CN 105695556A
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- inoculation
- inoculation method
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- exponential model
- spiral
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- 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
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/24—Methods of sampling, or inoculating or spreading a sample; Methods of physically isolating an intact microorganisms
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- 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
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
- C12Q1/06—Quantitative determination
Abstract
The invention relates to the technical field of colony count and provides a screw inoculation method. The method includes: inoculating a sample according to a diminishing ratio by taking the Archimedes screw as an inoculation path; enabling logarithm or other types of liquid inoculation by the aid of rotary cams and rotating plates; after inoculation, distributing bacterial colonies on the screw path in a more and more dispersed manner along with radius increase; and counting the bacterial colonies on a plate from the outer periphery to the center by the aid of grids for counting according to inoculation rules, so that the concentration of microbes in the sample is obtained. The screw inoculation method which is quick, accurate and uniform is mainly used for microbe counting, mutagenesis analysis, drug sensitivity analysis and single colony separation, is capable of saving materials and reducing cost for laboratories and especially saving energy and time and is higher in accuracy and repeatability as compared with a traditional plate method.
Description
Technical field
The present invention relates to a kind of colony counting technical field, particularly relate to a kind of spiral inoculation method。
Background technology
Spiral inoculation method for counting colonies is a kind of new food, cosmetics and the Rapid Quantification of microorganism in research sample。This spiral inoculation method for counting colonies is created in 1976, by Donnelly, C.B., J.E.Gilchrist et al. proposes at first, through the practical application of more than 20 years, external microorganism educational circles is generally approved, the method being listed as international analysis chemist association (AOAC), is included as national standard by U.S. food Drug Administration (FDA)。
Summary of the invention
The present invention provides a kind of spiral inoculation method, make it be applied to microorganism count, mutagenesis analysis, Analysis of Drug Susceptibility separate with single bacterium colony, can saving consumptive material for laboratory, reduce cost, it is often more important that be sparing of one's energy and the time, result relatively classic flat-plate method is more accurate, repeatability is better。
In order to solve the problems referred to above, the present invention provides a kind of spiral inoculation method, wherein, comprises the following steps:
S10, sample are inoculation track according to Archimedian screw, with a ratio inoculation sample successively decreased；
S20, by rotating cam and flap, liquid can be made to reach the inoculation of logarithmic or alternate manner；
After S30, inoculation, bacterium colony is distributed on helical trajectory, and is distributed increasingly to disperse with the increase of radius；
S40, utilization are by inoculating the grid that rule carries out counting, and from flat board, the bacterium colony on plate is counted by outer circumference central authorities；
The concentration of microorganism in sample is obtained after S50, calculating。
Preferably, described bacterium colony refers under certain condition, such as aerobic situation, nutritional condition, pH, cultivation temperature and time, the total number of bacterial colonies that every gram or every milliliter sample grows out。
Preferably, described spiral inoculation method includes exponential model, and exponential model refers to that, in seeded process, sample size is successively decreased with exponential model, separates for colony counting and single bacterium colony, wherein original bacterium solution concentration range: 4.0 × 10 in Rotating Plates method2-4.0×105Cfu/ml。
Preferably, described spiral inoculation method includes slow exponential model, slow exponential model refers in seeded process, sample size is successively decreased with exponential model, separates for colony counting and single bacterium colony in Rotating Plates method, and speed is slightly slow, when using wet flat board, select slow exponential model, it is prevented that sample is outer under the effect of centripetal force to be spattered, wherein original bacterium solution concentration range: 4.0 × 102-4.0×105Cfu/ml。
Preferably, described spiral inoculation method includes ratio mode, and in ratio mode, inoculum concentration is successively decreased to the air line distance at center is proportional, the scope of inoculum concentration is narrower, contributes to amplifying bacterial number reaction to antibiotic or mutagenic agent in only small gradient scope。
Preferably, described spiral inoculation method includes homogeneous pattern, and homogeneous pattern refers in rotary course, and sample is with constant speed inoculation。
Preferably, described spiral inoculation method includes lawn pattern, and lawn pattern refers to and inoculates with constant rate of speed according to homogeneous pattern, but closeer than homogeneous pattern one times of track, homogeneous, intensive growth can be produced on whole flat board。
Preferably, described spiral inoculation method includes circulation pattern, and circulation pattern refers to simulation exponential model, but inoculation track is not a helix, but a series of circle, the inoculum density often organizing circle is identical。
Compared with prior art, the invention have the advantages that
The present invention is a kind of inoculation method quick, accurate, uniform, be mainly used in microorganism count, mutagenesis analysis, Analysis of Drug Susceptibility separate with single bacterium colony, consumptive material can be saved for laboratory, reduce cost, the more important thing is and be sparing of one's energy and the time, result relatively classic flat-plate method is more accurate, repeatability is better。
Detailed description of the invention
In order to make the purpose of the present invention, technical scheme and advantage clearly understand, below the present invention is described in further detail, but example is not as a limitation of the invention。
Embodiments of the invention include a kind of spiral inoculation method, wherein, comprise the following steps:
S10, sample are inoculation track according to Archimedian screw, with a ratio inoculation sample successively decreased；
S20, by rotating cam and flap, liquid can be made to reach the inoculation of logarithmic or alternate manner；
After S30, inoculation, bacterium colony is distributed on helical trajectory, and is distributed increasingly to disperse with the increase of radius；
S40, utilization are by inoculating the grid that rule carries out counting, and from flat board, the bacterium colony on plate is counted by outer circumference central authorities；
The concentration of microorganism in sample is obtained after S50, calculating。
Bacterium colony refers under certain condition, such as aerobic situation, nutritional condition, pH, cultivation temperature and time, the total number of bacterial colonies that every gram or every milliliter sample grows out。
In the present embodiment, spiral inocalation method includes following pattern:
Exponential model (inoculum concentration: 50 μ l):
In seeded process, sample size is successively decreased with exponential model。Rotating Plates method separates for colony counting and single bacterium colony。Original bacterium solution concentration range: 4.0 × 102-4.0×105Cfu/ml
Exponential model inoculation method is different from traditional pouring-type inoculation method, eliminate the tedious steps of continuous gradient dilution, the usage quantity of the consumptive material such as culture dish, culture medium 3/4 during pattern detection can be reduced, save the plenty of time of experimenter, thus obtaining being widely recognized as of the departments such as quality inspection, agriculture inspection and commodity inspection simultaneously。
Exponential model inocalation method can also be used in spiral gradient terminal test and SGE test。Utilize spiralplater to inoculate antibiotic, form the radially continuous Concentraton gradient of antibiotic。Bacterium solution is radially coated in culture medium, multiple strain can be tested simultaneously。Antibiotic concentration is gradually increased to center by edge, and different strain can be formed different inhibitory action, utilizes professional software to can determine whether this kind of antibiotic minimum inhibitory concentration MIC for this strain。
Slow exponential model (inoculum concentration: 50 μ l): inoculum concentration, pattern are identical with exponential model, and speed is slightly slow。When using wet flat board, select slow exponential model, it is prevented that sample is outer under the effect of centripetal force to be spattered。Original bacterium solution concentration range: 4.0 × 102-4.0×105Cfu/ml
Ratio mode (inoculum concentration: 20 μ l): inoculum concentration is successively decreased to the air line distance at center is proportional, and therefore ratio index pattern is much narrower for the scope of inoculum concentration。This inoculation pattern contributes to amplifying bacterial number reaction to antibiotic or mutagenic agent in only small gradient scope。
Homogeneous pattern (inoculum concentration: 20 μ l): in rotary course, sample is with constant speed inoculation。Can be used for inoculation Salmonella in Ames test Ames, the interaction of two kinds of microorganisms of research, utilize rarer original bacterium solution to obtain a large amount of single bacterium colonies etc.。
Lawn pattern (inoculum concentration: 20 μ l): inoculate with constant rate of speed according to homogeneous pattern, but closeer than homogeneous pattern one times of track, can produce homogeneous, intensive growth on whole flat board。This inoculation pattern is in culture medium, and coated sample uniform, accurate, such as the research for KB scraps of paper antibiotics susceptibility test or virus plaque。
Circulation pattern (inoculum concentration: 50 μ l): simulation exponential model, but inoculation track is not a helix, but a series of circle, the inoculum density often organizing circle is identical, mainly more convenient during counting。
Above inoculum concentration is all as the criterion with 90mm culture dish。
Described above to the disclosed embodiments, makes professional and technical personnel in the field be capable of or uses the present invention。The multiple amendment of these embodiments be will be apparent from for those skilled in the art, and generic principles defined herein can without departing from the spirit or scope of the present invention, realize in other embodiments。Therefore, the present invention is not intended to be limited to the embodiments shown herein, and is to fit to the widest scope consistent with principles disclosed herein and features of novelty。
Claims (8)
1. a spiral inoculation method, it is characterised in that comprise the following steps:
S10, sample are inoculation track according to Archimedian screw, with a ratio inoculation sample successively decreased；
S20, by rotating cam and flap, liquid can be made to reach the inoculation of logarithmic or alternate manner；
After S30, inoculation, bacterium colony is distributed on helical trajectory, and is distributed increasingly to disperse with the increase of radius；
S40, utilization are by inoculating the grid that rule carries out counting, and from flat board, the bacterium colony on plate is counted by outer circumference central authorities；
The concentration of microorganism in sample is obtained after S50, calculating。
2. spiral inoculation method as claimed in claim 1, it is characterised in that described bacterium colony refers under certain condition, such as aerobic situation, nutritional condition, pH, cultivation temperature and time, the total number of bacterial colonies that every gram or every milliliter sample grows out。
3. spiral inoculation method as claimed in claim 1, it is characterized in that, described spiral inoculation method includes exponential model, exponential model refers in seeded process, sample size is successively decreased with exponential model, Rotating Plates method separates for colony counting and single bacterium colony, wherein original bacterium solution concentration range: 4.0 × 102-4.0×105Cfu/ml。
4. spiral inoculation method as claimed in claim 1, it is characterized in that, described spiral inoculation method includes slow exponential model, and slow exponential model refers to that, in seeded process, sample size is successively decreased with exponential model, Rotating Plates method separates for colony counting and single bacterium colony, speed is slightly slow, when using wet flat board, selects slow exponential model, spatter outside preventing sample under the effect of centripetal force, wherein original bacterium solution concentration range: 4.0 × 102-4.0×105Cfu/ml。
5. spiral inoculation method as claimed in claim 1, it is characterized in that, described spiral inoculation method includes ratio mode, in ratio mode, inoculum concentration is successively decreased to the air line distance at center is proportional, the scope of inoculum concentration is narrower, contributes to amplifying bacterial number reaction to antibiotic or mutagenic agent in only small gradient scope。
6. spiral inoculation method as claimed in claim 1, it is characterised in that described spiral inoculation method includes homogeneous pattern, and homogeneous pattern refers in rotary course, sample is with constant speed inoculation。
7. spiral inoculation method as claimed in claim 1, it is characterized in that, described spiral inoculation method includes lawn pattern, and lawn pattern refers to and inoculates with constant rate of speed according to homogeneous pattern, but closeer than homogeneous pattern one times of track, can produce homogeneous, intensive growth on whole flat board。
8. spiral inoculation method as claimed in claim 1, it is characterised in that described spiral inoculation method includes circulation pattern, circulation pattern refers to simulation exponential model, but inoculation track is not a helix, but a series of circle, the inoculum density often organizing circle is identical。
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| CN201610090903.2A CN105695556A (en) | 2016-02-18 | 2016-02-18 | Screw inoculation method |
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| CN201610090903.2A CN105695556A (en) | 2016-02-18 | 2016-02-18 | Screw inoculation method |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107870224A (en) * | 2017-11-03 | 2018-04-03 | 安徽省农业科学院烟草研究所 | A kind of field identification method for evaluating corn aspergillus flavus infection resistance ability |
| CN110564809A (en) * | 2019-09-18 | 2019-12-13 | 武汉迪艾斯科技有限公司 | rotary streaking inoculation method |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104593473A (en) * | 2013-10-31 | 2015-05-06 | 大连大公环境检测有限公司 | Method for detecting bacterial microbes in soil |
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2016
- 2016-02-18 CN CN201610090903.2A patent/CN105695556A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104593473A (en) * | 2013-10-31 | 2015-05-06 | 大连大公环境检测有限公司 | Method for detecting bacterial microbes in soil |
Non-Patent Citations (2)
| Title |
|---|
| 北京五洲东方科技发展有限公司: "快速实现菌落总数计数的好方法", 《中国教育装备采购网》 * |
| 北京五洲东方科技发展有限公司: "快速实现菌落总数计数的好方法", 《生物在线》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107870224A (en) * | 2017-11-03 | 2018-04-03 | 安徽省农业科学院烟草研究所 | A kind of field identification method for evaluating corn aspergillus flavus infection resistance ability |
| CN110564809A (en) * | 2019-09-18 | 2019-12-13 | 武汉迪艾斯科技有限公司 | rotary streaking inoculation method |
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Application publication date: 20160622 |
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