CN103540656A - Jellyfish species identification method - Google Patents
Jellyfish species identification method Download PDFInfo
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- CN103540656A CN103540656A CN201310221007.1A CN201310221007A CN103540656A CN 103540656 A CN103540656 A CN 103540656A CN 201310221007 A CN201310221007 A CN 201310221007A CN 103540656 A CN103540656 A CN 103540656A
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Abstract
The invention relates to the technical field of biology and provides a jellyfish species identification method. The jellyfish species identification method comprises the following steps: firstly extracting jellyfish DNA (deoxyribonucleic acid), and performing jellyfish 16s RNA (ribonucleic acid) database analysis on the jellyfish DNA; then designing a 16s RNA specific primer and a Taqman probe according to a 16s RNA homology analysis result; next, taking the jellyfish DNA as a template and utilizing the specific primer to perform Taqman probe detection so as to obtain curve data; and finally determining the species of jellyfish according to an amplification result. According to the jellyfish species identification method provided by the invention, the species of the jellyfish can be determined according to the jellyfish DNA, and the species identification can be performed against adult individuals and jellyfish larvae with incomplete shapes. Simultaneously, the jellyfish 16s RNA specific primer is designed according to the 16s RNA homology analysis result, the targeted property is strong and the accuracy is high.
Description
Technical field
The present invention relates to biological technical field, be specifically related to biological species discrimination method.
Background technology
The nearly more than ten years, the littoral jellyfish of China occurs in a large number, to the marine eco-environment, has brought certain impact.Complete adult individuality is than being easier in the discriminating of kind in form for jellyfish, but in form, the kind discrimination ratio of incomplete adult individuality and the jellyfish young is more difficult.
Summary of the invention
The object of the invention is to, provide a kind of species of jellyfish discrimination method, to address the above problem.
Technical problem solved by the invention can realize by the following technical solutions:
Species of jellyfish discrimination method, is characterized in that, first, utilizes clustalx software to jellyfish 16s RNA database analysis; Then, according to 16s RNA homology analysis result, utilize primer analysis software design jellyfish 16s RNA Auele Specific Primer and Taqman probe; Then, extract jellyfish DNA, take jellyfish DNA as template, utilize Auele Specific Primer to carry out Taqman probe in detecting to obtain curve data; Finally, determine species of jellyfish.The present invention determines the kind of jellyfish according to the DNA of jellyfish, can carry out Identification of Species for incomplete adult individuality and the jellyfish young in form.Simultaneously the present invention is according to 16s RNA homology analysis consequence devised jellyfish 16s RNA Auele Specific Primer and Taqman probe, with strong points, accuracy rate is high.
By jellyfish DNA extraction liquid, extract jellyfish DNA.When (the CTAB method after utilizing us to improve) used, directly 100-200g sample is put into jellyfish DNA extraction liquid, utilized jellyfish DNA extraction liquid to carry out jellyfish DNA extraction.Prior art, normally carries out freezing preservation by the sample of collection, and the present invention is directly kept at sample in extracting solution, and the operation after having avoided, has reached time saving and energy saving effect.
Use real time PCR instrument to adopt Real-Time Fluorescent Quantitative PCR Technique to detect jellyfish DNA, determine species of jellyfish.The present invention adopts technique scheme, both can use manpower and material resources sparingly, and can simplify production stage again.Real time PCR(be again real-time quantitative PCR) refer to during PCR index amplification and change and immediately measure specificity product by continuous monitoring fluorescent signal, do not need to take out PCR product and carry out separation.Real-Time Fluorescent Quantitative PCR Technique has not only been realized the leap of PCR from qualitative to quantitative, and compares with conventional PCR, and it has specificity and more by force, effectively solves PCR pollution problem, level of automation high.
The utensil using in experimental implementation process is all sterile-processed.The reagent using is DNase free level, to avoid jellyfish DNA degradation.
Because 98% of jellyfish is moisture, process not in time very easily degraded.Therefore, jellyfish sample collection, jellyfish DNA extraction interval in half an hour with interior the best.
Present method can be bitten for the sea of sand, aurelia, Australia spot jellyfish, jellyfish carry out kind discriminating.
Accompanying drawing explanation
Fig. 1 is schema of the present invention;
Fig. 2 is the result figure that the sea of sand is bitten pattern detection;
Fig. 3 is the result figure of aurelia pattern detection;
Fig. 4 is the result figure of Australia spot pattern detection;
Fig. 5 is the result figure of jellyfish pattern detection.
Embodiment
For technique means, creation characteristic that the present invention is realized, reach object and effect is easy to understand, below in conjunction with concrete diagram, further set forth the present invention.
With reference to Fig. 1, Fig. 2, Fig. 3, Fig. 4 and Fig. 5.Species of jellyfish discrimination method, first, extracts jellyfish DNA, and jellyfish DNA is carried out to jellyfish 16s RNA database analysis; Then, according to 16s RNA, with originality analytical results, design jellyfish 16s RNA Auele Specific Primer; Then, take jellyfish DNA as template, utilize Auele Specific Primer to carry out Taqman probe in detecting to obtain amplification curve; Finally, according to amplification, determine species of jellyfish.The present invention determines the kind of jellyfish according to the DNA of jellyfish, can carry out Identification of Species for incomplete adult individuality and the jellyfish young in form.Simultaneously the present invention is according to 16s RNA homology analysis consequence devised jellyfish 16s RNA Auele Specific Primer Taqman probe, with strong points, accuracy rate is high.
By jellyfish DNA extraction liquid, extract jellyfish DNA.When (the CTAB method after our improvement) used, directly 100-200g sample is put into jellyfish DNA extraction liquid, utilized jellyfish DNA extraction liquid to carry out jellyfish DNA extraction.Prior art, normally carries out freezing preservation by the sample of collection, and the present invention is directly kept at sample in extracting solution, and the operation after having avoided, has reached time saving and energy saving effect.
In aforesaid operations step, part operation step is less demanding to working order, also can synchronously carry out.As a kind of preferred version, in Fig. 1, first carry out jellyfish sample collecting, carry out jellyfish DNA extraction again, carry out at the same time jellyfish information acquisition, design Auele Specific Primer and Taqman probe, finally take jellyfish DNA as template, utilize Auele Specific Primer to carry out Taqman probe in detecting.
Use real time PCR instrument to adopt Real-Time Fluorescent Quantitative PCR Technique to detect jellyfish DNA, determine species of jellyfish.The present invention adopts technique scheme, both can use manpower and material resources sparingly, and can simplify production stage again.Real time PCR(be again real-time quantitative PCR) refer to during PCR index amplification and change and immediately measure specificity product by continuous monitoring fluorescent signal, do not need to take out PCR product and carry out separation.Real-Time Fluorescent Quantitative PCR Technique has not only been realized the leap of PCR from qualitative to quantitative, and compares with conventional PCR, and it has specificity and more by force, effectively solves PCR pollution problem, level of automation high.
The utensil using in experimental implementation process is all sterile-processed.The reagent using is DNase free level, to avoid jellyfish DNA degradation.
Because 98% of jellyfish is moisture, process not in time very easily degraded.Therefore, jellyfish sample collection, jellyfish DNA extraction interval in half an hour with interior the best.Present method can be bitten for the sea of sand, aurelia, Australia spot jellyfish, jellyfish carry out kind discriminating.In Fig. 2, Fig. 3, Fig. 4 and Fig. 5, A, B are that the sea of sand is bitten; C, D are aurelia; E, F are Australia spot jellyfish; G, H are jellyfish.
More than show and described ultimate principle of the present invention and principal character and advantage of the present invention.The technician of the industry should understand; the present invention is not restricted to the described embodiments; that in above-described embodiment and specification sheets, describes just illustrates principle of the present invention; without departing from the spirit and scope of the present invention; the present invention also has various changes and modifications, and these changes and improvements all fall in the claimed scope of the invention.The claimed scope of the present invention is defined by appending claims and equivalent thereof.
Claims (5)
1. species of jellyfish discrimination method, is characterized in that, first, extracts jellyfish DNA, and jellyfish DNA is carried out to jellyfish 16s RNA database analysis; Then, according to 16s RNA, with originality analytical results, design jellyfish 16s RNA Auele Specific Primer and Taqman probe; Then, take jellyfish DNA as template, utilize Auele Specific Primer to carry out Taqman probe in detecting to obtain curve data; Finally, according to amplification, determine species of jellyfish.
2. species of jellyfish discrimination method according to claim 1, is characterized in that: by jellyfish DNA extraction liquid, extract jellyfish DNA.(the CTAB method after our improvement).
3. species of jellyfish discrimination method according to claim 2, is characterized in that: during use, directly 100-200g sample is put into jellyfish DNA extraction liquid, utilized jellyfish DNA extraction liquid to carry out jellyfish DNA extraction.
4. species of jellyfish discrimination method according to claim 1, is characterized in that: use real time PCR instrument to adopt Real-Time Fluorescent Quantitative PCR Technique to detect jellyfish DNA, determine species of jellyfish.
5. according to the species of jellyfish discrimination method described in any one in claim 1-4, it is characterized in that: present method can be bitten for the sea of sand, aurelia, Australia spot jellyfish, jellyfish carry out kind discriminating.
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| CN201310221007.1A CN103540656A (en) | 2013-06-04 | 2013-06-04 | Jellyfish species identification method |
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| CN201310221007.1A CN103540656A (en) | 2013-06-04 | 2013-06-04 | Jellyfish species identification method |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104593489A (en) * | 2014-12-26 | 2015-05-06 | 中国科学院烟台海岸带研究所 | Specific primer for rapidly detecting white cyanea nozakii and detection method of specific primer |
| CN105331712A (en) * | 2015-11-19 | 2016-02-17 | 大连工业大学 | Method for rapidly identifying species of salted rhopilema esculentum kishinouye |
| KR20160073625A (en) * | 2014-12-17 | 2016-06-27 | 한국해양과학기술원 | Primer for species identification of the nine kinds of jellyfish from 28S ribosomal DNA origin |
-
2013
- 2013-06-04 CN CN201310221007.1A patent/CN103540656A/en active Pending
Non-Patent Citations (2)
| Title |
|---|
| KEITH M. BAYHA等: "A new Taqman_ PCR-based method for the detection and identification of scyphozoan jellyfish polyps", 《HYDROBIOLOGIA》 * |
| 王建艳等: "基于mt－16S rDNA 和mt－COI 基因的海月水母分子生物学鉴定方法和检测技术", 《应用生态学报》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20160073625A (en) * | 2014-12-17 | 2016-06-27 | 한국해양과학기술원 | Primer for species identification of the nine kinds of jellyfish from 28S ribosomal DNA origin |
| KR101693616B1 (en) | 2014-12-17 | 2017-01-06 | 한국해양과학기술원 | Primer for species identification of the nine kinds of jellyfish from 28S ribosomal DNA origin |
| CN104593489A (en) * | 2014-12-26 | 2015-05-06 | 中国科学院烟台海岸带研究所 | Specific primer for rapidly detecting white cyanea nozakii and detection method of specific primer |
| CN105331712A (en) * | 2015-11-19 | 2016-02-17 | 大连工业大学 | Method for rapidly identifying species of salted rhopilema esculentum kishinouye |
| CN105331712B (en) * | 2015-11-19 | 2019-02-12 | 大连工业大学 | A kind of method that Rapid identification pickling silk floss bites kind |
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Application publication date: 20140129 |