CN107228928A - The detection method of soil microbial biomass - Google Patents
The detection method of soil microbial biomass Download PDFInfo
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- CN107228928A CN107228928A CN201710408112.4A CN201710408112A CN107228928A CN 107228928 A CN107228928 A CN 107228928A CN 201710408112 A CN201710408112 A CN 201710408112A CN 107228928 A CN107228928 A CN 107228928A
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Abstract
The invention discloses a kind of detection method of soil microbial biomass, including：Take after soil sample progress sterilization treatment to be measured, moisture is dried in baking oven, and detect the content of organic matter；Microorganism fungus kind is added into soil sample to be measured, and is uniformly mixed；The crop seed of budding is planted in the soil sample to be measured added with microorganism fungus kind, and cultivated；The pedotheque after culture is taken, living bacteria count is determined.Using the detection method of the above-mentioned soil microbial biomass of the present invention, the microorganism fungus kind being analysed to is colonized in the native soil after processing, and simulate normal crop-planting environment, microorganism fungus kind is set to be bred with crop syntrophism, finally according to the situation of plantation culture, sample the microorganism living bacteria count amount of detection, can dynamic of the measure specified microorganisms in soil of accurate qualitative, quantitative colonize and expand numerous amount result.
Description
Technical field
The present invention relates to technical field of biological, more particularly to a kind of detection method of soil microbial biomass.
Background technology
Soil microbial biomass refers to volume in soil and is less than 5 × 103μm biological total amount, but plant such as root system of plant living
Etc. being not included, it is soil organic matter fractions living.Although soil microbial biomass only accounts for 3% of the soil organism or so,
But it is the driving force of plant food conversion, organic carbon metablism and contaminant degradation, is had in soil fertility and the ecosystem
Play the role of important.By detecting soil microbial biomass, whole microbiologic population or some of special in soil can be known
Population status, further learns the situation of change of farmland ecosystem, and Soil Microorganism activity and to organic matter residuum
The speed and intensity of decomposition.
At present for the detection method of soil microbial biomass, mainly have：Direct Microscopical Method For Detection (colony counting method), substrate for induction
Breathing Method, componential analysis, stifling cultivation, stifling extraction method.
Wherein, stifling extraction method be it is most popular it is a kind of determine soil microbial biomass method, procedure be by
Soil to be measured is after pharmacy fumigation, and Soil Microorganism is killed, and killed microbial body is newly added micro- life of people's original soil sample
Thing decomposes (mineralising) and releases CO2, according to the CO discharged2Amount and microbial body mineralization rate constant Kc can calculate the soil sample
Carbon amounts in microorganism, the size of carbon amounts just reflects the size of microbial biomass.Process can be known from the above, determine only
Only it is a macroscopical amount, and microorganism property and state can not be embodied.And another colony counting method process is direct sample
Micro- sem observation is used afterwards, and the more original backwardness of method is not illustrated herein.
And stifling cultivation is commonly used to determine microbial biomass --- the carbon in soil contaminated by crude oil, this method is by soil moisture shape
Condition influence is larger, to Free Ca CO3The high soil of content, flooded soils, pH<4.5 soil and applied recently organic fertilizer or
The soil of green manure, its measurement result is unreliable, is relatively limited using scope.And weak point is that the consuming time is long, is unsuitable for
Soil sample soil microbial biomass is air-dried to determine.
Componential analysis is to estimate that soil microbial biomass contains according to the content of the specific biological metabolism composition of certain in soil
Amount, the weak point of the method is to extract ATP efficiency and result is inaccurate, by different soils and different phosphorus contents influenceed conversion coefficient and
The influence of measure causes accidental error very big, and costly continuous mode is extremely complex simultaneously for required reagent.
The method of above-mentioned various detections, on the one hand the microbial biomass of detection is the micro- life of static soil departing from plant growth
Object amount；On the other hand detection be in soil all overall microorganism fungus kind syntrophisms to state when detecting.And can not be accurate
Dynamic of the measure microorganism of qualitative, quantitative in soil colonizes the numerous amount result of expansion, and can not be accurate to a certain specific micro- life
The growth and breeding state of thing.
The content of the invention
It is a primary object of the present invention to provide a kind of detection method of soil microbial biomass, it is intended to accurate qualitative, quantitative
Determine dynamic of the specified microorganisms in soil and colonize the numerous amount result of expansion.
To achieve the above object, the detection method of soil microbial biomass of the invention, comprises the following steps：
Take after soil sample progress sterilization treatment to be measured, moisture is dried in baking oven, and detect the content of organic matter；
Microorganism fungus kind is added into the soil sample to be measured, and is uniformly mixed；
The crop seed of budding is planted in the soil sample to be measured added with microorganism fungus kind, and cultivated；
The pedotheque after culture is taken, living bacteria count is determined.
Using the detection method of the above-mentioned soil microbial biomass of the present invention, the microorganism fungus kind being analysed to is colonized in processing
In native soil afterwards, and normal crop-planting environment is simulated, microorganism fungus kind is bred with crop syntrophism, finally according to
Plant the situation of culture, sample the microorganism living bacteria count amount of detection, can accurate qualitative, quantitative measure specified microorganisms
Dynamic in soil colonizes the numerous amount result of expansion.
Brief description of the drawings
Below in conjunction with drawings and Examples, the invention will be further described, in accompanying drawing：
Fig. 1 is that dynamic of the microorganism of detection of the embodiment of the present invention in soil expands numerous curve map.
Embodiment
The realization, functional characteristics and advantage of the object of the invention will be described further in conjunction with the embodiments.It should be appreciated that this
The specific embodiment of place description is not intended to limit the present invention only to explain the present invention.
The present invention, which is provided, a kind of can detect soil microbial biomass that microorganism colonizing in soil expands numerous growing state
Detection method, comprises the following steps：
S10, takes after soil sample progress sterilization treatment to be measured, moisture is dried in baking oven, and detect the content of organic matter；
S20, microorganism fungus kind is added into soil sample to be measured, and uniformly mix；Wherein, the amount of microorganism fungus kind is organic matter
3% (weight) of content；
S30, the crop seed sprouted is planted in the soil sample to be measured added with microorganism fungus kind, and is cultivated；
S40, takes the pedotheque after culture, determines living bacteria count.
The above method process of the present invention, is that will need the extraordinary microorganism fungus kind of analysis, colonize in soil sample, then most
Bigization simulation planting environment, makes microorganism fungus kind be bred with crop syntrophism, finally according to the situation of plantation culture, sampling
The microorganism living bacteria count amount of detection, analysis microorganism colonizes the numerous growing state of expansion in soil, depicts microorganism and exists
Growth curve in soil；The quantitative a certain microorganism of certainty can thus be obtained and colonize the numerous amount of expansion in soil.
Step S10 is obtained after pedotheque, is first calculated the content of soil with organic matter, on the one hand can tentatively be sentenced
Disconnected soil fertility situation, on the other hand can be for microorganism wt in estimation soil sample, and the weight of general microorganism, which accounts for soil, to be had
3% or so of machine matter weight, the quantization reference that can be used for during subsequently colonizing.Meanwhile, first by soil to be measured in the step
Earth sample carries out sterilization treatment, and sterilization treatment purpose is to colonize Reverse transcriptase/interference of microorganism to reduce, and is also simultaneously
In order to lift the accuracy of subsequent detection.Certainly, the mode of pedotheque sterilization treatment is preferred to use high pressure steaming in step S10
The mode of vapour sterilizing (121 DEG C, 0.1MPa, 30min) is carried out, and while completing, can not influence the crumb structure of soil sample, have
Beneficial to the soil texture for keeping pedotheque script.Afterwards, moisture is dried in 60 DEG C~80 DEG C baking ovens, water is on the one hand removed
Divide the accuracy of the organic quality detection of interference effect, be on the other hand conducive to preventing other miscellaneous bacterias infection in soil before the planting
Growth, microorganism fungus kind colonizing and detecting in influence this case.And temperature control is in this 60 DEG C~80 DEG C appropriate scope
Section, further can avoid high temperature from causing physical property to change, and destroy the physical arrangement and composition of soil.
Then step S20, the microbial biomass obtained according to budget, it would be desirable to after the microorganism fungus kind of detection and sterilization treatment
Soil sample mixing colonize, be then transferred in plant pot.In this step, the addition of microorganism fungus kind is according to big with estimation
Suitable organic matter weight 3% is caused to be added.
Wherein, when step S20 is implemented, microorganism fungus kind to be detected can be a kind of or two or more mixes
Close.If using single culture, then this method detection is that the single culture colonizes the numerous amount situation of expansion in soil；
If two or more mixed bacterias, then what is finally detected is the numerous amount situation of expansion of symbiosis.
During plantation, soil is irrigated with sterilized water further with the crop-planting sprouted in plant pot by step S30.
Certainly before plantation starts, the soil sample in plant pot can be taken, microorganism living bacteria count in soil sample is determined with colony counting method,
The bacterium number now determined, is referred to as follow-up blank control.During culture, according to the strain colonized it is different, with
And the difference for the crop planted, correspondence is during crop cultivation is planted, and giving suitable growth conditions makes it normally give birth to
It is long.
Finally during culture is planted, plant pot Soil K+adsorption microorganism fungus kind number of viable therein is regularly taken.
Wherein, take to detect the soil of microorganism fungus kind in step S40, preferably taken under conditions of root system is not hindered as far as possible close
The soil of root system, around crop root, is influenceed, the breeding of strain is with crop by edaphon with root respiration
More preferably, the result of detection is better for the relevance of growth.Meanwhile, based on the situation of microorganism fungus kind growth metabolism, fetch earth earth
The time of sample detection is used in the range of potted plant growth starts 7 days~35 days.The basic place of microorganism fungus kind growth before 7 days
The development stage is also in extremely low degree and crop seed, the result of sampling and measuring substantially compares no practical significance；35 days
After, the microorganism fungus kind around crop root is basicly stable, has substantially reached that microorganism colonizes the stage of ripeness of breeding.
After detection is completed, microorganism living bacteria count amount will be detected, analysis microorganism colonizes the numerous growing state of expansion in soil, retouched
Draw growth curve of the microorganism in soil.
Wherein, it is to carry out sterilization treatment to original soil sample to be measured to disappear by implementation in above-mentioned steps and implementation Process
Except interference, if the microorganism fungus kind to be colonized is the special strain of colonial morphology such as bacillusmusilaginosiengineering, or
By strain to be colonized isotope marks culture, the convenience and accuracy of subsequent detection can be strengthened, while follow-up
In the case of crop cultivation detection, eugonic, it is convenient to omit this link of the pedotheque inactivation treatment.
Using the detection method of the above-mentioned soil microbial biomass of the present invention, the microorganism fungus kind being analysed to is colonized in processing
In native soil afterwards, and normal crop-planting environment is simulated, microorganism fungus kind is bred with crop syntrophism, finally according to
Plant the situation of culture, sample the microorganism living bacteria count amount of detection, can accurate qualitative, quantitative measure specified microorganisms
Dynamic in soil colonizes the numerous amount result of expansion.
The details of detection method to make above-mentioned soil microbial biomass of the invention is more conducive to the understanding of those skilled in the art
And implementation, and the progressive effect of this case is protruded, lifted below by way of specific embodiment come the above to this case
Example explanation.
Embodiment 1
Prepare before S11, experiment：Randomly select soil sample gauze (more than 4 layers) parcels tight, outer layer newspaper or brown paper
Wrap, be positioned in autoclave sterilization pot and sterilize, while preparation running water and various utensils are placed in autoclave after wrapping
Sterilizing, autoclave is set as 121 DEG C, 0.1MPa, 30min.After sterilizing terminates, each article is cooled to room temperature；
S12, the soil sample after sterilizing is placed in baking oven and dries moisture, and oven temperature sets 60 DEG C.Soil sampling is surveyed after drying
Determine the content of organic matter.
S20, starts experiment：The addition ratio to pedotheque microorganism is calculated according to the content meter of organic matter in soil sample first
Example, account for the content of organic matter by microorganism 3% is calculated, and the soil sample for weighing 4 parts of equivalent weights is stand-by；Then organic matter in soil sample is pressed
Content 3% weigh 4 parts of the microbial bacterial agent (bacillus subtilis) of equivalent weight, 4 parts of microbial bacterial agents are added to 4 one by one
In part soil sample, it is uniformly mixed, is finally fitted into standby in plant pot；
S30, plant pot soil is irrigated with sterilized water, and the wheat seed sprouted is put in basin center, is placed in 25 DEG C of illumination
Start to plant culture experiment in constant incubator；
S40, microorganism colonizes the numerous tracking of expansion：The soil sampling before the wheat culture in potted plant starts, is surveyed with colony counting method
Determine microorganism living bacteria count in soil sample, in triplicate, compared as blank soil sample；Pot experiment started after 7 days, as far as possible
Do not hinder the pedotheque taken under conditions of root system close to root system, microorganism living bacteria count is determined with colony counting method, repeat three
It is secondary；By that analogy, pot experiment starts 10 days, 14 days, 17 days, 21 days, 24 days, 28 days, 31 days and 35 days, and soil sampling is surveyed respectively
Determine living bacteria count.During soil sampling, potted plant culture terminates experiment after 35 days, takes root soil to determine living bacteria count.
S50, data analysis：According to the microorganism living bacteria count amount of sampling detection, microorganism determining in soil is analyzed
The numerous growing state of expansion is grown, growth curve of the microorganism in soil is depicted.
Embodiment 2
In the embodiment 2, according to the identical step of embodiment 1, bacillusmusilaginosiengineering is colonized, it is same in potted plant culture
Root soil is taken to determine living bacteria count.
The potted plant of the microorganism fungus kind of above example 1-2 detections colonizes the numerous data result such as following table of expansion, according to above-mentioned inspection
The growth curve that the data of survey are described is as shown in Figure 1.
| Time (d) | Bacillus subtilis bacterial content (million/gram) | Bacillusmusilaginosiengineering content (million/gram) |
| 0 | 1 | 1.1 |
| 7 | 1.8 | 1.9 |
| 14 | 2.3 | 2.5 |
| 21 | 3.6 | 4 |
| 28 | 4.7 | 4.9 |
| 31 | 5 | 5.3 |
| 35 | 5.2 | 5.4 |
The preferred embodiments of the present invention are these are only, are not intended to limit the scope of the invention, it is every to utilize this hair
Equivalent structure or equivalent flow conversion that bright description is made, or directly or indirectly it is used in other related technology necks
Domain, is included within the scope of the present invention.
Claims (8)
1. a kind of detection method of soil microbial biomass, it is characterised in that comprise the following steps：
Take after soil sample progress sterilization treatment to be measured, moisture is dried in baking oven, and detect the content of organic matter；
Microorganism fungus kind is added into the soil sample to be measured, and is uniformly mixed；
The crop seed of budding is planted in the soil sample to be measured added with microorganism fungus kind, and cultivated；
The pedotheque after culture is taken, living bacteria count is determined.
2. the detection method of soil microbial biomass as claimed in claim 1, it is characterised in that take the soil sample to be measured to be gone out
After bacterium processing, moisture is dried in baking oven, and is detected in content of organic matter step,
The sterilization treatment is high pressure steam sterilization.
3. the detection method of soil microbial biomass as claimed in claim 1 or 2, it is characterised in that take the soil sample to be measured to enter
After row sterilization treatment, moisture is dried in baking oven, and is detected in content of organic matter step,
60 DEG C~80 DEG C of temperature control in the baking oven.
4. the detection method of soil microbial biomass as claimed in claim 1 or 2, it is characterised in that into the soil sample to be measured
Add in microorganism fungus kind, and uniform blend step,
The microorganism fungus kind is added by the 3% of the content of organic matter of detection.
5. the detection method of soil microbial biomass as claimed in claim 1 or 2, it is characterised in that into the soil sample to be measured
Add in microorganism fungus kind, and uniform blend step,
The microorganism fungus kind is single culture or mixed bacteria.
6. the detection method of soil microbial biomass as claimed in claim 1 or 2, it is characterised in that into the soil sample to be measured
Add in microorganism fungus kind, and uniform blend step,
The microorganism fungus kind is the strain with isotope marks.
7. the detection method of soil microbial biomass as claimed in claim 1 or 2, it is characterised in that take the soil after culture
Earth sample, is determined in living bacteria count step,
The pedotheque is the soil close to crop root.
8. the detection method of soil microbial biomass as claimed in claim 1 or 2, it is characterised in that by the crop of the budding
Seed is planted in the soil sample to be measured added with microorganism fungus kind, and is carried out in incubation step, and the time of the culture is 7
My god~35 days.
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Cited By (2)
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| CN110771292A (en) * | 2019-11-14 | 2020-02-11 | 神华宝日希勒能源有限公司 | Ecological maintenance-free method suitable for vegetation in strip mine refuse dump |
| CN113444612A (en) * | 2021-05-20 | 2021-09-28 | 北京工业大学 | Microcosmic culture device and application thereof in quantitative analysis of soil carbon diffusion and microorganism utilization process |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110771292A (en) * | 2019-11-14 | 2020-02-11 | 神华宝日希勒能源有限公司 | Ecological maintenance-free method suitable for vegetation in strip mine refuse dump |
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Application publication date: 20171003 |