CN107868760A - A kind of cultural method of Rhodopseudomonas palustris - Google Patents
A kind of cultural method of Rhodopseudomonas palustris Download PDFInfo
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- CN107868760A CN107868760A CN201611249071.0A CN201611249071A CN107868760A CN 107868760 A CN107868760 A CN 107868760A CN 201611249071 A CN201611249071 A CN 201611249071A CN 107868760 A CN107868760 A CN 107868760A
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- 241000190950 Rhodopseudomonas palustris Species 0.000 title claims abstract description 30
- 238000000034 method Methods 0.000 title claims abstract description 23
- 241000894006 Bacteria Species 0.000 claims abstract description 33
- 230000001580 bacterial effect Effects 0.000 claims abstract description 22
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 10
- 239000002609 medium Substances 0.000 claims description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 6
- 244000068988 Glycine max Species 0.000 claims description 5
- 235000010469 Glycine max Nutrition 0.000 claims description 5
- 239000001888 Peptone Substances 0.000 claims description 5
- 108010080698 Peptones Proteins 0.000 claims description 5
- 239000012530 fluid Substances 0.000 claims description 5
- 239000001963 growth medium Substances 0.000 claims description 5
- 235000015097 nutrients Nutrition 0.000 claims description 5
- 235000019319 peptone Nutrition 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 5
- 239000011780 sodium chloride Substances 0.000 claims description 5
- 229910001220 stainless steel Inorganic materials 0.000 claims description 5
- 239000010935 stainless steel Substances 0.000 claims description 5
- 230000001954 sterilising effect Effects 0.000 claims description 5
- 238000003756 stirring Methods 0.000 claims description 5
- 239000012137 tryptone Substances 0.000 claims description 5
- 238000007707 calorimetry Methods 0.000 claims description 4
- 238000012613 in situ experiment Methods 0.000 claims description 4
- 239000012533 medium component Substances 0.000 claims description 2
- 241000208340 Araliaceae Species 0.000 claims 1
- 235000005035 Panax pseudoginseng ssp. pseudoginseng Nutrition 0.000 claims 1
- 235000003140 Panax quinquefolius Nutrition 0.000 claims 1
- 235000008434 ginseng Nutrition 0.000 claims 1
- 238000005286 illumination Methods 0.000 abstract description 14
- 238000002474 experimental method Methods 0.000 abstract description 8
- 238000004519 manufacturing process Methods 0.000 abstract description 7
- 230000000694 effects Effects 0.000 abstract description 3
- 238000011065 in-situ storage Methods 0.000 abstract description 3
- 230000009286 beneficial effect Effects 0.000 abstract description 2
- 210000004369 blood Anatomy 0.000 abstract description 2
- 239000008280 blood Substances 0.000 abstract description 2
- 230000002503 metabolic effect Effects 0.000 abstract description 2
- 210000001916 photosynthetic cell Anatomy 0.000 abstract description 2
- 230000035945 sensitivity Effects 0.000 abstract description 2
- 238000012546 transfer Methods 0.000 abstract description 2
- 210000004027 cell Anatomy 0.000 description 12
- 230000000243 photosynthetic effect Effects 0.000 description 10
- 239000012153 distilled water Substances 0.000 description 5
- 210000000601 blood cell Anatomy 0.000 description 4
- 239000000049 pigment Substances 0.000 description 4
- 230000005526 G1 to G0 transition Effects 0.000 description 3
- 230000007423 decrease Effects 0.000 description 3
- 241000190932 Rhodopseudomonas Species 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 241000589516 Pseudomonas Species 0.000 description 1
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 238000009360 aquaculture Methods 0.000 description 1
- 244000144974 aquaculture Species 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 239000010842 industrial wastewater Substances 0.000 description 1
- 238000009630 liquid culture Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
- 230000004962 physiological condition Effects 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 229960005486 vaccine Drugs 0.000 description 1
- 238000004065 wastewater treatment Methods 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- Wood Science & Technology (AREA)
- Microbiology (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Tropical Medicine & Parasitology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The present invention relates to one kind to use the micro hot systems of light, passes through Rhodopseudomonas palustris cultural method that is high-precision, controlling illumination wavelength, power and temperature in high sensitivity.Strain is brought back to life first and transfers 3 on behalf of strain stoste, standard curve is made using blood plate counting method, according to this experiment with computing bacterium solution bacterial population, so as to ensure bacterial population in fixed range.Then by the micro hot systems of light, 532 nm wavelength light sources are opened, progress luminous power is 4 W/m2, temperature be 30 DEG C Rhodopseudomonas palustris growth in situ heat score-curve determine.The thermokinetic parameters and the quantity of heat production of different metabolic phase of exponential phase are finally calculated, determine light λ=532 nm, 4 W/m2The photosynthetic cells effect of bacterium can be most transferred, is metabolized beneficial to bacterial growth.The Rhodopseudomonas palustris lag phase time of this method culture is short, only with 27395 s;Exponential phase is up to 48.1851 μ W, 1343.66 J for up to 45570 s, thermal power and thermal discharge;Present invention illumination reaction condition simple to operate, bacterium is accurate, and reaction is efficient.
Description
Technical field
The present invention relates to a kind of cultural method of Rhodopseudomonas palustris, using light-micro hot systems, by high accuracy,
Wavelength, power and the temperature of illumination are controlled in high sensitivity, and bacterium is cultivated.Belong to field of microbial culture technology, have
Body is related to a kind of cultural method of Rhodopseudomonas palustris.
Background technology
The factor of photosynthetic bacterium growth is influenceed in addition to own physiological condition, also has some external factor, such as temperature, soda acid
Degree, the initial concentration of substrate and illumination can all limit the growth of bacterium.The wherein just energy source of photosynthetic bacterium growth, it is
The basic premise of photosynthetic bacteria existence.Photosynthetic pigments are pigments specific to photosynthetic organism, are to convert light energy into chemistry
The key substance of energy.Different photosynthetic bacteria strains contains different photosynthetic pigments, and the type and quantity of photosynthetic pigments are to light
Capture produces material impact, therefore growth of the light to photosynthetic bacteria is most important.
Rhodopseudomonas(Rhodopseudomonas)It is most commonly seen in photosynthetic bacteria and that there is Practical significance one
Individual category, Rhodopseudomonas palustris are its representative strain, Industrial Wastewater Treatment, hydrogen manufacturing, aquaculture, planting industry, cosmetics and
The fields such as health products have a wide range of applications, but are concentrated mainly on culture for Rhodopseudomonas palustris cultural method at present
In the improvement of base, such as cultivation temperature, acid-base value, substrate formula and concentration, for Rhodopseudomonas palustris culture illumination condition
The document report seen of research be the effect grown using complex light to it, and in single wavelength, certain power and temperature
Growth yet there are no document report under illumination.
The content of the invention
It is an object of the invention to the vacancy for existing area research, proposes a kind of culture side of Rhodopseudomonas palustris
Method.Present invention illumination reaction condition simple to operate, bacterium is accurate, and reaction is efficient.
To realize this purpose, the present invention uses light-micro hot systems, by controlling the wavelength of illumination and the marsh of power
Red pseudomonas cultural method.Strain is brought back to life first and 3 generations of transferring are as strain stoste, and standard is made using blood plate counting method
Curve determination, experiment with computing bacterium solution bacterial population according to this, so as to ensure bacterial population in fixed range.Then using light-micro heat system
System, 532 nm wavelength light sources are opened, progress luminous power is 4 W/m2Rhodopseudomonas palustris growth in situ heat score-curve survey
It is fixed.Finally by the thermokinetic parameters and the quantity of heat production of different metabolic phase for calculating exponential phase, monochromatic light λ=532 are determined
Nm, 4 W/m2The photosynthetic cells effect that power can most transfer bacterium makes bacterium keep good growth situation, beneficial to bacterial growth generation
Thank.The Rhodopseudomonas palustris lag phase time that this method is turned out is short, only with 27395 s;Exponential phase for up to
45570 s, thermal power and thermal discharge are up to 48.1851 μ W, 1343.66 J.
The method of the present invention includes step in detail below:
1st, the preparation of culture medium:Medium component is weighed respectively:The g of tryptone 15.0, the g of soya peptone 5.0, sodium chloride 5.0
G, distilled water is dissolved in, 1.0 L are settled to distilled water after stirring, it is 7.3 ± 0.2 to survey solution PH, is sub-packed in 250 mL cones
Shape bottle, after sealing, 121 DEG C of 30 min of sterilizing.
2nd, Rhodopseudomonas palustris actication of culture:Freeze-dried vaccine is forwarded to from slant medium 4-5 mL Liquid Cultures
Base, bring back to life strain.Transferred for 3 generations again as strain stoste.
3rd, Rhodopseudomonas palustris bacterial concentration:Counted using counting method of blood cell.Put down with the strain after resurrection
Plate culture, 30 DEG C are placed in, intensity of illumination is to be counted after cultivating 24-48 hours in 2500 lux illumination incubators, makes standard
Curve.Rhodopseudomonas palustris bacterial concentration measure measures its its OD value at 660 nm using spectrophotometric, passes through mark
Directrix curve fit equation determines bacterial population so that bacterium solution bacterial population used is about 10 every time8Individual/mL.
With this experiment with computing bacterium solution bacterial population.
4th, Rhodopseudomonas palustris culture:In light-micro hot systems, 1.5 mL culture mediums and 1.5 mL are separately added into
The fluid nutrient medium of Rhodopseudomonas palustris is inoculated with into sample cell and reference cell, then it is 15 mL's to be respectively charged into volume
In stainless steel sample cell and reference tube, constant temperature is to 30 DEG C, after baseline stability, opens 532 nm wavelength light sources, carries out light work(
Rate is 4 W/m2Rhodopseudomonas palustris growth in situ experiment, time 345600s.
The present invention has following features:
1st, the illumination of Rhodopseudomonas palustris bacterium of the present invention culture is specific wavelength monochromatic light, i.e. the nm of λ=532, and certain power is
4W/m2, specified temp be 30 DEG C be controlled by light-micro hot systems.
2nd, the condition of Rhodopseudomonas palustris of the present invention is constant temperature, and facultative aerobic culture can be by thin in incubation
" heat score-curve " of bacteria growing is monitored to its growth course state.And calculated after experiment terminates according to heat score-curve
The Rhodopseudomonas palustris lag phase time that this method is turned out is 27395 s;The exponential phase time is 45570 s, hot merit
Rate and thermal discharge are up to 48.1851 μ W, 1343.66 J respectively, in view of in production, come often through the time for extending logarithmic phase
The fermentation efficiency of raising bacterium is visible, and the Rhodopseudomonas palustris cultivated using the present invention has low cost, high efficiency, operation letter
Single, the accurate feature of illumination reaction condition of bacterium.
Brief description of the drawings
Fig. 1 is the type light of RD 496-CK 2000-micro hot systems reality that the present invention is used for Rhodopseudomonas palustris culture
Thing figure.
Fig. 2 is the heat production P- of the gained of the embodiment of the present invention 1tThe time of curve map and lower different growing stages, thermal change
Figure.
Fig. 3 is the heat production P- of the gained of the embodiment of the present invention 2tThe time of curve map and lower different growing stages, thermal change
Figure.
Fig. 4 is the heat production P- of the gained of the embodiment of the present invention 3tThe time of curve map and lower different growing stages, thermal change figure.
Embodiment
Technical scheme is further described below by way of specific embodiment.Following examples are to this hair
Bright further explanation, rather than limitation the scope of the present invention.
Embodiment 1
1)The preparation of culture medium:Weigh the g of tryptone 15.0, the g of soya peptone 5.0, the g of sodium chloride 5.0, distilled water 1.0
L, after stirring, its PH is surveyed as 7.3 ± 0.2,121 DEG C of 30 min of sterilizing.
2)Actication of culture:Bring back to life strain and 3 generations of transferring are as strain stoste.
3)Bacterial concentration:Counted using counting method of blood cell.With after resurrection strain carry out flat board culture, culture 24 ~
Count within 48 hours, making standard curve, experiment with computing bacterium solution bacterial population,.
4)Bacteria Culture:In light-Calorimetry system, be separately added into 1.5 mL culture mediums and 1.5 mL to be inoculated with marsh red
The fluid nutrient medium of pseudomonad is into sample cell and reference cell, then is respectively charged into the stainless steel sample cell that volume is 15 mL
In reference tube, constant temperature is to 30 DEG C, after baseline stability, opens 532 nm wavelength light sources, progress luminous power is 4 W/m2's
Rhodopseudomonas palustris growth in situ is tested, and the time is 345600 s.
Experiment terminate after according to heat score-curve, as shown in figure 1, A-B is lag phase, B-C exponential phases, C-D is
Stationary phase, D-E are decline phase.It is 27395 s to calculate the Rhodopseudomonas palustris lag phase time that this method is turned out;It is right
Number growth period is up to 48.1851 μ W, 1343.66 J respectively for up to 45570 s, thermal power and thermal discharge.
Embodiment 2
1)The preparation of culture medium:Weigh the g of tryptone 15.0, the g of soya peptone 5.0, the g of sodium chloride 5.0, distilled water 1.0
L, after stirring, its PH is surveyed as 7.3 ± 0.2,121 DEG C of 30 min of sterilizing.
2)Actication of culture:Bring back to life strain and 3 generations of transferring are as strain stoste.
3)Bacterial concentration:Counted using counting method of blood cell.With after resurrection strain carry out flat board culture, culture 24 ~
Count within 48 hours, make standard curve determination, experiment with computing bacterium solution bacterial population.
4)Bacteria Culture:In light-Calorimetry system, be separately added into 1.5 mL culture mediums and 1.5 mL to be inoculated with marsh red
The fluid nutrient medium of pseudomonad(Measure bacterium solution OD values before measure every time, bacterial population is determined by standard curve fit equation,
So that bacterium solution bacterial population used is about 10 every time8Individual/mL)Into sample cell and reference cell, then it is 15 mL to be respectively charged into volume
Stainless steel sample cell and reference tube in, set relevant parameter, constant temperature is to 30 DEG C, after baseline stability, open 532 nm
Wavelength light source, progress luminous power are 2 W/m2Rhodopseudomonas palustris growth in situ experiment, the time is 345600 s(96
h).
Test after terminating according to heat score-curve, as shown in Fig. 2 A'B' is lag phase, B'-C' exponential phases, C'-
D' is stationary phase, and D'-E' is decline phase.Calculating the Rhodopseudomonas palustris lag phase time that this method is turned out is
161200 s;The exponential phase time is 11362 s, and thermal power and thermal discharge are respectively 19.687 μ W, 135.00 J.Explanation
, illumination power is too small, and the growth metabolism of Rhodopseudomonas palustris is slack-off, and growth performance declines.
Embodiment 3
1)The preparation of culture medium:Weigh the g of tryptone 15.0, the g of soya peptone 5.0, the g of sodium chloride 5.0, distilled water 1.0
L, after stirring, its PH is surveyed as 7.3 ± 0.2,121 DEG C of 30 min of sterilizing.
2)Actication of culture:Bring back to life strain and 3 generations of transferring are as strain stoste.
3)Bacterial concentration:Counted using counting method of blood cell.With after resurrection strain carry out flat board culture, culture 24 ~
Count within 48 hours, make standard curve determination, experiment with computing bacterium solution bacterial population.
4)Bacteria Culture:In light-Calorimetry system, be separately added into 1.5 mL culture mediums and 1.5 mL to be inoculated with marsh red
The fluid nutrient medium of pseudomonad(Measure bacterium solution OD values before measure every time, bacterial population is determined by standard curve fit equation,
So that bacterium solution bacterial population used is about 10 every time8Individual/mL)Into sample cell and reference cell, then it is 15 mL to be respectively charged into volume
Stainless steel sample cell and reference tube in, set relevant parameter, constant temperature is to 30 DEG C, after baseline stability, open 532 nm
Wavelength light source, progress luminous power are 6 W/m2Rhodopseudomonas palustris growth in situ experiment, the time is 345600 s(96
h).
Test after terminating according to heat score-curve, as shown in figure 3, A 〞-B 〞 are lag phase, B 〞-C 〞 exponential phases, C 〞-D
〞 is stationary phase, and D 〞-E 〞 are decline phase.It is 142838 to calculate the Rhodopseudomonas palustris lag phase time that this method is turned out
s;The exponential phase time is 29549 s, and thermal power and thermal discharge are respectively 38.561 μ W, 842.66 J.Illustrate, illumination
Power is excessive, and the growth metabolism of Rhodopseudomonas palustris is equally slack-off, and growth performance declines.
Claims (2)
1. using the method for light-micro hot systems culture Rhodopseudomonas palustris, it is characterised in that high-precision, highly sensitive
Light specific wavelength is λ=532 nm, certain power 4W/m2, specified temp is 30 DEG C.
2. according to claim 1, a kind of cultural method of Rhodopseudomonas palustris, comprise the following steps:
1) preparation of culture medium:Medium component is the g of tryptone 15.0, the g of soya peptone 5.0, the g of sodium chloride 5.0, is distilled
The L of water 1.0, after stirring, its pH is surveyed as 7.3 ± 0.2,121 DEG C of 30 min of sterilizing;
2) actication of culture:Bring back to life strain and 3 generations of transferring are as strain stoste;
3) Bacteria Culture:In light-Calorimetry system, it is separately added into 1.5 mL culture mediums and 1.5 mL is inoculated with the red false list in marsh
The fluid nutrient medium of born of the same parents bacterium is into sample cell and reference cell, then is respectively charged into the stainless steel sample cell and ginseng that volume is 15 mL
Than in pipe, bacterium solution bacterial population used is about 108Individual/mL, constant temperature is to 30 DEG C, after baseline stability, opens 532 nm wavelength lights
Source, row luminous power are 4 W/m2Rhodopseudomonas palustris growth in situ experiment, the time is 345600 s.
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Cited By (1)
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CN112877242A (en) * | 2021-02-05 | 2021-06-01 | 广东工业大学 | Bacterial source photosensitizer for wastewater treatment and preparation method and application thereof |
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Cited By (1)
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CN112877242A (en) * | 2021-02-05 | 2021-06-01 | 广东工业大学 | Bacterial source photosensitizer for wastewater treatment and preparation method and application thereof |
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