CN110305942A - A kind of Fish Environment DNA moves water and returns become characteristic test method and device - Google Patents

A kind of Fish Environment DNA moves water and returns become characteristic test method and device Download PDF

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CN110305942A
CN110305942A CN201910653310.6A CN201910653310A CN110305942A CN 110305942 A CN110305942 A CN 110305942A CN 201910653310 A CN201910653310 A CN 201910653310A CN 110305942 A CN110305942 A CN 110305942A
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fish
environment dna
water
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characteristic test
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CN110305942B (en
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郭辉
杨文俊
罗玉兰
段文刚
滕素芬
黄明海
李利
黄卫
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Changjiang River Scientific Research Institute Changjiang Water Resources Commission
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Abstract

The present invention, which provides a kind of Fish Environment DNA and moves water, returns become characteristic test method and device, is related to water environment and Ecology protects field, the test method includes: step (1): acquisition target detection fish living body biological sample;Step (2): it moves water in Fish Environment DNA and returns in the characteristic test device that becomes and carry out Fish Environment DNA and fall off characteristic test;Step (3): on the basis of step (2), carry out the characteristic test of Fish Environment DNA degradation;Step (4): on the basis of step (2), carry out the test of Fish Environment DNA characterization of adsorption;Step (5): test analysis environment DNA concentration changes with time process, parsing Fish Environment DNA move water and return the characteristic that becomes.The present invention can the dynamic water of accurate quantification test different types and Life Stages Fish Environment DNA return the characteristic that becomes, there is important scientific application value.

Description

A kind of Fish Environment DNA moves water and returns become characteristic test method and device
Technical field
Water, which is moved, the present invention relates to water environment and Ecology protection field, in particular to a kind of Fish Environment DNA returns the characteristic that becomes Test method and device.
Background technique
Hydraulic engineering influences to be widely noticed on fish reproduction habitat, and monitoring and evaluation is the important foundation of protection.Environment DNA prison It surveys and is used as the high timeliness new methods of non-intruding, be widely used in and judge rare or invasive species presence or absence.Research finds environment DNA There are correlativities for concentration and piscine organism characteristic and water body environment characteristic, but attempt to be applied to forecast assessment fish and its Breeding habitat distribution, history of life behavior, biomass etc., it is still necessary to solve how to test environment DNA move water return become characteristic this Bottleneck problem.
Summary of the invention
Water is moved the purpose of the present invention is to overcome the shortcomings of background technique, providing a kind of Fish Environment DNA and returns the characteristic survey that becomes Method for testing and device.
To achieve the above object, the present invention, which provides a kind of Fish Environment DNA and moves water, returns the characteristic test method that becomes, and feature exists In, comprising the following steps:
A kind of Fish Environment DNA moves water and returns the characteristic test method that becomes, comprising the following steps:
Step (1): acquisition target detection fish living body biological sample;
Step (2): Fish Environment DNA move water return in the characteristic test device that becomes carry out Fish Environment DNA fall off characteristic reality It tests;
Step (3): on the basis of step (2), carry out the characteristic test of Fish Environment DNA degradation;
Step (4): on the basis of step (2), carry out the test of Fish Environment DNA characterization of adsorption;
Step (5): test analysis environment DNA concentration changes with time process, parsing Fish Environment DNA move water and return the spy that becomes Property.
Further, the step (1) acquires target detection fish living body biological sample from field or breeding field, including Adult fish, juvenile fish, fish-egg, while the basic biological nature of fish, including weight, size, stage of development are tested, it is fitted raising and train in pond Should for 24 hours, during which fasting.
Further, target detection fish living body biological sample is put into the dynamic water of Fish Environment DNA and returned by the step (2) Become in characteristic test device, (u, m/s) different in flow rate, water temperature (T, DEG C), dissolved oxygen (DO, mg/L) factor operating condition feelings is respectively set Scape, different time intervals (t in 2~4di, s) and take quantitative water sample (Vi, mL), respectively in the interior filter for passing through 0.22 μm of aperture for 24 hours Film vacuum filtration, saves backup under the conditions of filtered filter membrane is placed in -20 DEG C, makes for step (5) test environment DNA concentration With.
Further, the step (3) is on the basis of step (2), by target detection fish living body biological sample from The dynamic water of Fish Environment DNA, which is returned in the characteristic test device that becomes, to be taken out, different time intervals (t in 4~8di, s) and take quantitative water sample (Vi, mL), it is filtered by vacuum respectively in the interior filter membrane by 0.22 μm of aperture for 24 hours, filtered filter membrane is placed under the conditions of -20 DEG C It saves backup, is used for step (5) test environment DNA concentration.
Further, the step (4) is on the basis of step (2), by target detection fish living body biological sample from The dynamic water of Fish Environment DNA, which is returned in the characteristic test device that becomes, to be taken out, and a certain amount of suspension bed sediment particle is added, builds different turbidity water flows Operating condition scene, different time intervals (t in 1~2di, s) and take quantitative water sample (Vi, mL), pass through 0.22 μ of aperture interior for 24 hours respectively The filter membrane of m is filtered by vacuum, and saves backup under the conditions of filtered filter membrane is placed in -20 DEG C, tests environment DNA concentration for step 5 It uses.
Further, the step (5) is using quantitative instant Polymerase Chain Reaction (real-time qPCR) technology Environment DNA number of fragments (N in testing procedure (2), (3) and (4) in filtered filter membranei, copies), calculate environment DNA Concentration (Ci, copies/mL):
Ci=Ni/Vi
Fish Environment DNA moves water and returns the characteristic test device that becomes to can be considered as one to be thoroughly mixed water body, Fish Environment DNA Dynamic water returns the time-varying process for the characteristic that becomes to characterize using one-dimensional model:
Wherein CiFor environment DNA concentration, copies/mL;T is time, s;S is environment DNA expulsion rate, copies/ (mL d);k1For environment DNA degradation rate, 1/d;k2For environment DNA adsorption rate, 1/d.
When step (2) Mesichthyes environment DNA drops to stable state,
Therefore, (1) formula may be characterized as,
S=(k1+k2)C0 (2)
Return the characteristic test dress that becomes when target detection fish living body biological sample is moved water from Fish Environment DNA in step (3) After setting taking-up, S=0, then (1) formula be may be characterized as,
Therefore, Fish Environment DNA, which moves water time degradation process, may be characterized as following exponential model
Return the characteristic test dress that becomes when target detection fish living body biological sample is moved water from Fish Environment DNA in step (4) After setting taking-up, S=0, then (1) formula may be characterized as,
Therefore, Fish Environment DNA, which moves water time adsorption process, may be characterized as following exponential model
More than, C0Return the characteristic test device that becomes for target detection fish living body biological sample is moved water from Fish Environment DNA Initial steady state environment DNA concentration, copies/mL after middle taking-up;
Different time intervals (t in plot step (2), (3) and (4) respectivelyi, s) and environment DNA concentration (Ci, copies/mL) Scatter plot, analyzing water body environment DNA concentration changes over time rule during falling off, degrading and adsorbing etc. and return and, uses The homing methods such as weighted least-squares are fitted exponential model shown in formula (4) and (6), determine Fish Environment DNA expulsion rate (S), drop Solution rate (k1) and adsorption rate (k2) etc. keys return the characterisitic parameter that becomes.
A kind of Fish Environment DNA moves water and returns the characteristic test device that becomes, including for fish habitat annular channel, be located at annular Fish is blocked in the frequency conversion disk current generating system of runner side, the upstream fish screen for being set to frequency conversion disk current generating system upstream and downstream and downstream Grid, the Water Factors monitoring system positioned at the annular channel other side, ADV current meter and sampling system, the frequency conversion disk are made Streaming system includes variable-frequency motor, transmission belt, rotation axis and disk disc, the variable-frequency motor by transmission belt drive rotation axis into And disk disc is driven to rotate.
Further, the arrangement width of the disk disc and annular channel similar width, spacing is set as between disc 0.5~2cm.
Further, the annular channel is made of organic glass, and the upstream fish screen uses 0.5~2cm of diameter Honeycomb structure is made in round tube, and downstream fish screen is made of chemical fibre mesh sheet, and mesh size should be able to prevent test object fish from passing through.
Further, the upper end of the annular channel is equipped with locating support, the Water Factors monitoring system and ADV stream Fast instrument is erected on locating support, and the Water Factors monitoring system includes water temperature sensor, dissolved oxygen sensor, pH value biography Sensor, conductivity sensor, turbidity transducer.
The invention has the following beneficial effects:
1, Fish Environment DNA is returned the mechanism that becomes to be divided into fall off, degrade and adsorb three canonical process by the present invention, respectively in fish Class environment DNA moves water and returns development experiment in the characteristic Special testing device that becomes, and is based on qPCR technical testing different time nodes fish Environment DNA concentration draws the scatter plot of different time intervals environment DNA concentration, regression fit environment DNA concentration-time process Line, Fish Environment DNA concentration changes over time rule under conditions of quantitative analysis hydrodynamic force and Environmental Factors.
2, the Special testing device controllable the present invention is based on hydrodynamic force and other environmental factors, can quantitative analysis environmental factor The influence that water returns the characteristic that becomes is moved to Fish Environment DNA.
3, the present invention is based on the test and regression analysis of Fish Environment DNA concentration time course data, fish is quantitatively calculated The keys such as class environment DNA expulsion rate, degradation rate and adsorption rate return the characterisitic parameter that becomes.
Detailed description of the invention
Fig. 1 is the flow chart that Fish Environment DNA of the present invention moves that water returns the one of embodiment of the characteristic test method that becomes;
Fig. 2 is that the dynamic water of Fish Environment DNA of the present invention returns the planar structure of the one of embodiment of the characteristic test device that becomes to show It is intended to;
Fig. 3 is A-A cross section structure schematic diagram in Fig. 2;
Fig. 4 is B-B the schematic diagram of the section structure in Fig. 2;
Fig. 5 is that fish fall off environment DNA concentration-time process schematic;
Fig. 6 is that environment DNA concentration-time process schematic is degraded or adsorbed to fish.
In figure: 1- annular channel, 2- variable-frequency motor, 3- transmission belt, 4- rotation axis, 5- disk disc, the upstream 6- fish screen, The downstream 7- fish screen, 8- locating support, 9- environmental factor monitor system, 10-ADV current meter, 11- sampling system, 12- sampling Mouthful, 13- sampling valve.
Specific embodiment
Present pre-ferred embodiments are elaborated with reference to the accompanying drawing, but they are not constituted to limit of the invention It is fixed, it only illustrates, while the contents of the present invention and advantage are more clearly understood convenient for those skilled in the art.
It moves water as shown in Figure 1, the embodiment of the present invention provides a kind of Fish Environment DNA and returns the characteristic test method that becomes, feature It is, comprising the following steps:
Step (1): acquisition target detection fish living body biological sample;
Specifically, acquiring target detection fish living body biological sample, including adult fish, juvenile fish, fish-egg from field or breeding field Deng;Basic biological nature, including weight, size, stage of development of fish etc. are tested simultaneously.It is adapted in pond for 24 hours raising and train, during which Fasting.
Step (2): carry out Fish Environment DNA and fall off characteristic test;
Return the characteristic that becomes specifically, target detection fish living body biological sample is respectively put into multiple Fish Environment DNA and moves water In test device, (u, m/s) operating condition scene different in flow rate is set, such as 0.2,0.4,0.8,1.2m/s etc., test water temperature (T, DEG C), dissolved oxygen (DO, mg/L) factor.Different time intervals (t in 2~4di) take quantitative water sample (Vi, mL) and 50mL, such as 0h, 2h, 6h, 12h, 18h, for 24 hours, 1.5d, 2d, 3d, 4d etc..It will quantitative water sample (Vi, mL) and pass through 0.22 μm of aperture interior for 24 hours respectively Filter membrane vacuum filtration, saved backup under the conditions of filtered filter membrane is placed in -20 DEG C, for step (5) test environment DNA concentration It uses
Step (3): on the basis of step (2), carry out the characteristic test of Fish Environment DNA degradation;
Specifically, target detection fish living body biological sample is moved water from Fish Environment DNA on the basis of step (2) Return in the characteristic test device that becomes and takes out, different time intervals (t in 4~8di, s) and take quantitative water sample (Vi, mL) and 50mL, such as 0h, 2h, 6h, 12h, for 24 hours, 1.5d, 2d, 4d, 6d, 8d etc..It will quantitative water sample (Vi) pass through 0.22 μm of aperture in for 24 hours respectively Filter membrane vacuum filtration, saves backup under the conditions of filtered filter membrane is placed in -20 DEG C, makes for step (5) test environment DNA concentration With.
Step (4): on the basis of step (2), carry out the test of Fish Environment DNA characterization of adsorption;
Specifically, target detection fish living body biological sample is moved water from Fish Environment DNA on the basis of step (2) Return in the characteristic test device that becomes and take out, a certain amount of suspension bed sediment particle is added, builds different turbidity (Tur, NTU) flow conditions, 1 Different time intervals (t in~2di, s) and take quantitative water sample (Vi, mL) 50mL, such as 0min, 2min, 5min, 10min, 20min, 30min, 1h, 2h, 6h, 1d, 2d etc..It will quantitative water sample (Vi, mL) respectively in the interior filter membrane for passing through 0.22 μm of aperture for 24 hours Vacuum filtration, saves backup under the conditions of filtered filter membrane is placed in -20 DEG C, uses for step (5) test environment DNA concentration.
Fig. 2, Fig. 3 and Fig. 4 are please referred to, the Fish Environment DNA used in the step (2), (3) and (4) moves water and returns the spy that becomes System safety testing device, including the annular channel 1 for fish habitat, the frequency conversion disk current generating system positioned at 1 side of annular channel, setting In the upstream fish screen 6 and downstream fish screen 7, the water ring positioned at 1 other side of annular channel of frequency conversion disk current generating system upstream and downstream The border factor monitors system 9, ADV current meter 10 and sampling system 11, and the frequency conversion disk current generating system includes variable-frequency motor 2, passes Dynamic band 3, rotation axis 4 and disk disc 5, the variable-frequency motor 2 drive rotation axis 4 to drive disk disc 5 by transmission belt 3 Rotation, the disk disc 5 are formed by stacking up and down by multilayer disc piece, arrangement width and the 1 width phase of annular channel of disk disc 5 Closely, spacing may be configured as 0.5~2cm according to intensity of flow demand is made between disc, can avoid broken to the machinery of environment DNA segment It is bad.Water Factors monitor system 9 include water temperature (T, DEG C) sensor, dissolved oxygen (DO, mg/L) sensor and turbidity (Tur, The sensors such as NTU), ADV current meter 10 is for measuring water volume flow rate (u, m/s) and Turbulent characteristics, and sampling system 11 is for taking Different time intervals water sample.
Wherein, the annular channel 1 can be used the transparent materials such as organic glass and be made, facilitate observation fish movement state with And carry out environmental factor regulation;Its shape can be needed to be provided in round according to test or ellipse.
The upstream fish screen 6 can be used diameter 0.5~2cm round tube and honeycomb structure be made, and have both flow-shape adjustment function Energy;Downstream fish screen 7 is made of chemical fibre mesh sheet, and mesh size should be able to prevent test object fish from passing through.
As shown in figure 3, the upper end of the annular channel 1 is equipped with locating support 8, the Water Factors monitor 9 frame of system It is located on locating support 8, including the Water Factors sensor such as water temperature, dissolved oxygen, pH value, conductivity, turbidity, for monitoring water Body environmental factor change procedure.
The ADV current meter 10 is also erected on locating support 8, is convenient for measuring water body different spatial flow velocity and turbulent fluctuation Characteristic.
The sampling system 11 needs that multiple sample taps 12 are arranged along the depth of water according to test, and sampling valve is arranged in sample tap 12 13, take water sample in different time intervals, using quantitative PCR technique test environment DNA concentration, according to environment DNA concentration when Between changing rule, parsing Fish Environment DNA dynamic water return the characteristic that becomes.
Step (5): test analysis environment DNA concentration changes with time process, parsing Fish Environment DNA move water and return the spy that becomes Property.
The step (5) uses the instant Polymerase Chain Reaction of real-time quantitative (real-time qPCR) technical testing step (2), the environment DNA number of fragments (N in filter membrane has been filtered in (3) and (4)i), calculate environment DNA concentration.
Ci=Ni/Vi
Fish Environment DNA moves water and returns the characteristic test device that becomes to can be considered as one to be thoroughly mixed water body, Fish Environment DNA Dynamic water returns the time-varying process for the characteristic that becomes to characterize using one-dimensional model.
Wherein CiFor environment DNA concentration, copies/mL;T is time, s;S is environment DNA expulsion rate, copies/ (mL d);k1For environment DNA degradation rate, 1/d;k2For environment DNA adsorption rate, 1/d.
When step (2) Mesichthyes environment DNA drops to stable state,
Therefore,
S=(k1+k2)C0
Return the characteristic test dress that becomes when target detection fish living body biological sample is moved water from Fish Environment DNA in step (3) After setting taking-up, S=0, then one-dimensional model be may be characterized as,
Therefore, Fish Environment DNA, which moves water time degradation process, may be characterized as following exponential model
Return the characteristic test dress that becomes when target detection fish living body biological sample is moved water from Fish Environment DNA in step (4) After setting taking-up, S=0, then one-dimensional model may be characterized as,
Therefore, Fish Environment DNA, which moves water time adsorption process, may be characterized as following exponential model
More than, C0Return the characteristic test device that becomes for target detection fish living body biological sample is moved water from Fish Environment DNA Initial steady state environment DNA concentration, copies/mL after middle taking-up.
With reference to Fig. 5, plot step (2) different time intervals (ti, s) and fish fall off environment DNA concentration (Ci, copies/mL) Scatter plot, analyzing water body environment DNA concentration changes over time rule during falling off.
With reference to Fig. 6, plot step (3) or (4) different time intervals (ti, s) and fish fall off environment DNA concentration (Ci, Copies/mL scatter plot), analyzing water body environment DNA concentration change over time rule during degradation or absorption etc. return and. Intend Fish Environment DNA degradation and adsorption exponent model using homing methods such as weighted least-squares, determines that Fish Environment DNA falls off Rate (S), degradation rate (k1) or adsorption rate (k2) etc. keys return the characterisitic parameter that becomes, i.e. k1Or k1+k2=0.8011 1/d, C0= 1.8751×104Copies/mL, S=1.5021 × 104copies/(mL·d)。
Fish Environment DNA is returned the mechanism that becomes to be divided into fall off, degrade and adsorb three canonical process by the present invention, respectively in fish Environment DNA moves water and returns development experiment in the characteristic Special testing device that becomes, and is based on qPCR technical testing different time nodes fish ring Border DNA concentration, the scatter plot of drafting different time intervals environment DNA concentration, regression fit environment DNA concentration-time graph, Fish Environment DNA concentration changes over time rule under conditions of quantitative analysis hydrodynamic force and Environmental Factors.
The above description is merely a specific embodiment, but scope of protection of the present invention is not limited thereto, any Belong to those skilled in the art in the technical scope disclosed by the present invention, any changes or substitutions that can be easily thought of, all answers It is included within the scope of the present invention.

Claims (10)

1. a kind of Fish Environment DNA, which moves water, returns the characteristic test method that becomes, it is characterised in that: the following steps are included:
Step (1): acquisition target detection fish living body biological sample;
Step (2): it moves water in Fish Environment DNA and returns in the characteristic test device that becomes and carry out Fish Environment DNA and fall off characteristic test;
Step (3): on the basis of step (2), carry out the characteristic test of Fish Environment DNA degradation;
Step (4): on the basis of step (2), carry out the test of Fish Environment DNA characterization of adsorption;
Step (5): test analysis environment DNA concentration changes with time process, parsing Fish Environment DNA move water and return the characteristic that becomes.
2. a kind of Fish Environment DNA according to claim 1, which moves water, returns the characteristic test method that becomes, it is characterised in that: described The step of (1) from field or breeding field acquire target detection fish living body biological sample, including adult fish, juvenile fish, fish-egg, survey simultaneously The basic biological nature of fish, including weight, size, stage of development are tried, is adapted in pond for 24 hours raising and train, during which fasting.
3. a kind of Fish Environment DNA according to claim 1, which moves water, returns the characteristic test method that becomes, it is characterised in that: described The step of (2) by target detection fish living body biological sample be put into Fish Environment DNA move water return in the characteristic test device that becomes, respectively (u, m/s) different in flow rate, water temperature (T, DEG C), dissolved oxygen (DO, mg/L) factor operating condition scene are set, in 2~4d between different time Every (ti, s) and take quantitative water sample (Vi, mL), it is filtered by vacuum respectively in the interior filter membrane by 0.22 μm of aperture for 24 hours, it will be filtered Filter membrane saves backup under the conditions of being placed in -20 DEG C, uses for step (5) test environment DNA concentration.
4. a kind of Fish Environment DNA according to claim 1, which moves water, returns the characteristic test method that becomes, it is characterised in that: described The step of (3) on the basis of step (2), target detection fish living body biological sample from Fish Environment DNA moved into water return and become special It is taken out in system safety testing device, different time intervals (t in 4~8di, s) and take quantitative water sample (Vi, mL), pass through respectively interior for 24 hours The filter membrane vacuum filtration that 0.22 μm of aperture, saves backup under the conditions of filtered filter membrane is placed in -20 DEG C, tests for step (5) Environment DNA concentration uses.
5. a kind of Fish Environment DNA according to claim 1, which moves water, returns the characteristic test method that becomes, it is characterised in that: described The step of (4) on the basis of step (2), target detection fish living body biological sample from Fish Environment DNA moved into water return and become special It is taken out in system safety testing device, a certain amount of suspension bed sediment particle is added, build different turbidity water flow operating condition scenes, difference in 1~2d Time interval (ti, s) and take quantitative water sample (Vi, mL), it is filtered by vacuum respectively in the interior filter membrane by 0.22 μm of aperture for 24 hours, incited somebody to action The filter membrane filtered saves backup under the conditions of being placed in -20 DEG C, uses for step 5 test environment DNA concentration.
6. a kind of Fish Environment DNA according to claim 1, which moves water, returns the characteristic test method that becomes, it is characterised in that: described The step of (5) using in quantitative instant Polymerase Chain Reaction (real-time qPCR) technical testing step (2), (3) and (4) Environment DNA number of fragments (N in filtered filter membranei, copies), calculate environment DNA concentration (Ci, copies/mL):
Ci=Ni/Vi
Fish Environment DNA moves water and returns the characteristic test device that becomes to can be considered as one to be thoroughly mixed water body, and Fish Environment DNA moves water The time-varying process for the characteristic that becomes is returned to characterize using one-dimensional model:
Wherein CiFor environment DNA concentration, copies/mL;T is time, s;S is environment DNA expulsion rate, copies/ (mLd);k1 For environment DNA degradation rate, 1/d;k2For environment DNA adsorption rate, 1/d.
When step (2) Mesichthyes environment DNA drops to stable state,
Therefore, (1) formula may be characterized as,
S=(k1+k2)C0 (2)
Return in the characteristic test device that becomes when target detection fish living body biological sample is moved water from Fish Environment DNA in step (3) After taking-up, S=0, then (1) formula be may be characterized as,
Therefore, Fish Environment DNA, which moves water time degradation process, may be characterized as following exponential model
Return in the characteristic test device that becomes when target detection fish living body biological sample is moved water from Fish Environment DNA in step (4) After taking-up, S=0, then (1) formula may be characterized as,
Therefore, Fish Environment DNA, which moves water time adsorption process, may be characterized as following exponential model
More than, C0It is taken out to return in the characteristic test device that becomes target detection fish living body biological sample from the dynamic water of Fish Environment DNA Initial steady state environment DNA concentration afterwards, copies/mL;
Different time intervals (t in plot step (2), (3) and (4) respectivelyi, s) and environment DNA concentration (Ci, copies/mL) dissipate Point diagram, analyzing water body environment DNA concentration changes over time rule during falling off, degrading and adsorbing etc. and return and, using weighting The homing methods such as least square are fitted exponential model shown in formula (4) and (6), determine Fish Environment DNA expulsion rate (S), degradation rate (k1) and adsorption rate (k2) etc. keys return the characterisitic parameter that becomes.
7. a kind of Fish Environment DNA, which moves water, returns the characteristic test device that becomes, it is characterised in that: including the annular flow for fish habitat Fish is blocked in road, the frequency conversion disk current generating system positioned at annular channel side, the upstream that is set to frequency conversion disk current generating system upstream and downstream Grid and downstream fish screen, Water Factors monitoring system, ADV current meter and sampling system positioned at the annular channel other side, institute Stating frequency conversion disk current generating system includes variable-frequency motor, transmission belt, rotation axis and disk disc, and the variable-frequency motor passes through transmission belt It drives rotation axis and then drives the rotation of disk disc.
8. Fish Environment DNA as claimed in claim 7, which moves water, returns the characteristic test device that becomes, it is characterised in that: the disk disk The arrangement width of piece and annular channel similar width, spacing is set as 0.5~2cm between disc.
9. Fish Environment DNA as claimed in claim 7, which moves water, returns the characteristic test device that becomes, it is characterised in that: the annular flow Road is made of organic glass, and honeycomb structure, downstream fish screen is made using 0.5~2cm of diameter round tube in the upstream fish screen It is made of chemical fibre mesh sheet, mesh size should be able to prevent test object fish from passing through.
10. Fish Environment DNA as claimed in claim 7, which moves water, returns the characteristic test device that becomes, it is characterised in that: the annular flow The upper end in road is equipped with locating support, and the Water Factors monitoring system and ADV current meter are erected on locating support, the water Environmental factor monitoring system includes water temperature sensor, dissolved oxygen sensor, pH sensor, conductivity sensor, turbidity sensing Device.
CN201910653310.6A 2019-07-19 2019-07-19 Method and device for testing DNA (deoxyribonucleic acid) dynamic water return-drive characteristic of fish environment Active CN110305942B (en)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20110281273A1 (en) * 2009-01-29 2011-11-17 Spiber Inc. Method of making dna tag
CN103461232A (en) * 2013-08-08 2013-12-25 中国水产科学研究院长江水产研究所 Testing device and testing method for testing environmental preference behavior of fish
CN107099595A (en) * 2017-05-12 2017-08-29 中国长江三峡集团公司中华鲟研究所 Fish natural propagation monitoring method based on environment DNA technology
CN210560495U (en) * 2019-07-19 2020-05-19 长江水利委员会长江科学院 Fish environment DNA moves water and returns to trend characteristic test device

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20110281273A1 (en) * 2009-01-29 2011-11-17 Spiber Inc. Method of making dna tag
CN103461232A (en) * 2013-08-08 2013-12-25 中国水产科学研究院长江水产研究所 Testing device and testing method for testing environmental preference behavior of fish
CN107099595A (en) * 2017-05-12 2017-08-29 中国长江三峡集团公司中华鲟研究所 Fish natural propagation monitoring method based on environment DNA technology
CN210560495U (en) * 2019-07-19 2020-05-19 长江水利委员会长江科学院 Fish environment DNA moves water and returns to trend characteristic test device

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
BRIAN KLITGAARD HANSEN等: "The sceptical optimist: challenges and perspectives for the application ofenvironmental DNA in marine fisheries" *
SATOSHI YAMAMOTO等: "Environmental DNA as a ‘Snapshot’ of Fish Distribution: ACase Study of Japanese Jack Mackerel in Maizuru Bay, Sea of Japan" *
TAAL LEVI等: "Environmental DNA for the enumeration and management of Pacific salmon" *
赵明等: "环境DNA在水域生态中的研究进展" *
陈炼等: "环境DNA metabarcoding及其在生态学研究中的应用" *

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