CN104962474B - A kind of cultural method of alga cells - Google Patents
A kind of cultural method of alga cells Download PDFInfo
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Abstract
The present invention discloses a kind of cultural method of alga cells, it is related to technical field of cell culture, for the phenomenon that generating bubbling being alleviated or avoided in the alga cells layer being attached on supporting layer, improves the cultured output of alga cells in application Immobilized culture technology culture alga cells.The cultural method of the alga cells includes: to carry out shortening processing to the first algal filament, obtains second algal filament of the length less than the first algal filament；Second algal filament is seeded on supporting layer, the alga cells layer being attached on supporting layer is obtained；Alga cells in second algal filament in alga cells layer are cultivated.The cultural method of alga cells provided by the invention is in the Immobilized culture of alga cells.
Description
Technical field
The present invention relates to technical field of cell culture more particularly to a kind of cultural methods of alga cells.
Background technique
In existing alga cells incubation, Immobilized culture technology is used, this is because Immobilized culture skill more
Art has many advantages, such as small occupied area, low energy consumption, low cost and high yield.
Using the detailed process of Immobilized culture technology culture alga cells are as follows: constituted alga cells to be cultivated
Raw material algal filament is seeded on the supporting layer such as plate or dry goods, and alga cells layer (alga cells layer is formed on supporting layer
Including cells of superficial layer and bottom cell), then the gas of culture medium and suitable algae cell growth is provided to the alga cells layer
Body environment and illumination condition can cultivate the alga cells in raw material algal filament.It is thin in the algae being attached on supporting layer
In born of the same parents' layer, since cells of superficial layer is located at alga cells layer surface, sufficient illumination and carbon dioxide gas supply can be obtained,
Therefore the growth of cells of superficial layer is more vigorous, the most of cultured output being capable of providing in alga cells culture.
However, in above-mentioned incubation, it is thin that alga cells by oxygen caused by photosynthesis can remain in algae
In born of the same parents' layer；In addition, can also not remained in alga cells layer by the carbon dioxide gas that above-mentioned photosynthesis consumes.These residuals
Gas lead to alga cells layer bulging and generate bubbling, which raises the cells of superficial layer in alga cells layer, causes surface layer
Cell is far from supporting layer, it is difficult to and culture medium is obtained from supporting layer, is affected so as to cause the growth of cells of superficial layer, it is final to cause
Making the cultured output of alga cells reduces.
Summary of the invention
The purpose of the present invention is to provide a kind of cultural methods of alga cells, for training in application Immobilized culture technology
When supporting alga cells, the phenomenon that generating bubbling is alleviated or avoided in the alga cells layer being attached on supporting layer, it is thin to improve algae
The cultured output of born of the same parents.
To achieve the goals above, the invention provides the following technical scheme:
The present invention provides a kind of cultural methods of alga cells comprising:
Shorten processing to first algal filament, obtains the second algal filament that length is less than first algal filament, described the
One algal filament and second algal filament are made of alga cells；
Second algal filament is seeded on the supporting layer, the alga cells layer being attached on the supporting layer is obtained；
Alga cells in second algal filament described in the alga cells layer are cultivated.
The present invention provides a kind of cultural methods of alga cells, carry out shortening processing in advance to the first algal filament, be grown
The second algal filament less than the first algal filament is spent, the second algal filament is seeded on supporting layer and forms alga cells layer.Due to the second algae
The length of silk is shorter, therefore in the alga cells layer, there are more gap between the second algal filament, these gaps are mutually interconnected
Lead to and the surface for the alga cells layer that goes directly, so that being produced during alga cells culture because of the photosynthesis of alga cells
The gases such as raw oxygen and the carbon dioxide provided to carry out alga cells culture to alga cells layer, can more hold
It changes places and is escaped from alga cells layer by above-mentioned gap, so that being not likely to produce bubbling in alga cells layer.Therefore, and now
Have in technology frequently to generate to be bubbled in alga cells layer and compare, the cultural method of alga cells provided by the invention can reduce or
It avoids the gases such as oxygen, carbon dioxide from accumulating caused bubbling phenomenon in alga cells layer, and then is alleviated or avoided by rousing
The problem of cells of superficial layer is raised caused by bubble finally improves algae so that cells of superficial layer is easy to obtain culture medium from supporting layer
The cultured output of cell.
In addition, in the present invention, since the second trichome length is shorter, having between the second algal filament in alga cells layer
More gap, so that the gases such as the illumination provided from the external world to alga cells layer and carbon dioxide are easy to stitch via these
Gap reaches the bottom cell in alga cells layer, so that bottom cell obtains the gas such as more illumination and carbon dioxide
Body further increases the cultured output of alga cells.
Detailed description of the invention
The drawings described herein are used to provide a further understanding of the present invention, constitutes a part of the invention, this hair
Bright illustrative embodiments and their description are used to explain the present invention, and are not constituted improper limitations of the present invention.In the accompanying drawings:
Fig. 1 is the flow chart of alga cells cultural method provided in an embodiment of the present invention；
Fig. 2 is the flow chart of the first improvement project of alga cells cultural method shown in FIG. 1；
Fig. 3 is the first specific flow chart of step 40 in alga cells cultural method shown in FIG. 1；
Fig. 4 is second of specific flow chart of step 40 in alga cells cultural method shown in FIG. 1；
Fig. 5 is the third specific flow chart of step 40 in alga cells cultural method shown in FIG. 1；
Fig. 6 is the 4th kind of specific flow chart of step 40 in alga cells cultural method shown in FIG. 1；
Fig. 7 is the specific flow chart for building immobilization cultivation platform.
Specific embodiment
The cultural method for the alga cells that embodiment provides in order to further illustrate the present invention, with reference to the accompanying drawings of the specification
It is described in detail.
Referring to Fig. 1, the cultural method of alga cells provided in an embodiment of the present invention includes:
Step 20 carries out the first algal filament to shorten processing, obtains second algal filament of the length less than the first algal filament, the first algal filament
It is made of with the second algal filament alga cells to be cultivated.Specifically, after determining the alga cells to be cultivated, the algae is obtained
The first algal filament that cell is constituted, first algal filament can be acquired from nature, can also be by carrying out to existing raw material algal filament
Small-scale Liquid Culture obtains, in addition, can also be by the resulting production of the cultural method of alga cells provided in an embodiment of the present invention
Object is as the first algal filament, to realize the Cyclic culture of alga cells, using the embodiment of the present invention to reduce the training of alga cells
Feeding process improves the breeding efficiency of alga cells.
Second algal filament is seeded on supporting layer by step 30, obtains the alga cells layer being attached on supporting layer.In the step
In rapid, it includes but is not limited to spray, scratch and filter etc. that the second algal filament, which is seeded in the method on supporting layer,.Wherein, it sprays
Inoculation method can be that the culture medium for being mixed with the second algal filament is sprayed on supporting layer so that the second algal filament is attached to supporting layer
On, this method has the advantages that easy to implement and cost is relatively low；The inoculation method of blade coating can be that will be mixed with the culture of the second algal filament
Base is applied on supporting layer, is fixed on the second algal filament on supporting layer by the mechanical force in knifing process, and this method, which has, to be connect
Kind advantage secured, not easily to fall off；The inoculation method of suction filtration can be that the second algal filament is placed on supporting layer, covers supporting layer
Bottle,suction bottleneck, and the portion gas in bottle,suction is extracted out by the side bifurcations on bottle,suction, keep the air pressure in bottle,suction low
In the air pressure outside bottle,suction, to be attached on supporting layer using the second algal filament of air pressure official post inside and outside bottle,suction, this method is not
Only the second algal filament can be made to be more firmly attached on supporting layer, additionally it is possible to it is thin to the algae in the second algal filament to mitigate seeded process
The active damage of born of the same parents.Those skilled in the art can be suitable according to specific cultivation needs, operation difficulty and cost, selection
Inoculation method.
Step 40 cultivates the alga cells in the second algal filament each in alga cells layer.Training for alga cells
It supports, it usually needs gases, culture medium, the illumination such as the carbon dioxide for being suitable for algae cell growth etc. are provided to alga cells layer, made
Algae cell growth (growth course of alga cells include cell itself volume become larger and the division growth of cell etc.),
In, when providing carbon dioxide gas to alga cells layer, carbon dioxide gas can be passed through culture medium according to certain ventilation ratio
In, being provided to alga cells layer together with culture medium, (ventilation ratio is represented when being passed through gas into liquid, is passed through in the unit time
The conventional unit of the volume of gas in liquid, the ratio with total liquid volume, ventilation ratio is vvm, and the ventilation ratio of 1vvm represents 1
The volume that the intracorporal gas of liquid of 1 unit volume is passed through in minute is 1 unit volume, i.e., the intracorporal gas of liquid is passed through in 1 minute
Volume it is equal with the total volume of liquid).So that alga cells thickness degree increases, it therefore, can be according to alga cells layer
Thickness determines the opportunity harvested to alga cells.It should be noted that removing alga cells itself, the culture institute of alga cells
The product of acquisition can also be the substances such as polysaccharide, protein and triglycerides of alga cells secretion in incubation.
The present invention provides a kind of cultural methods of alga cells, carry out shortening processing in advance to the first algal filament, be grown
The second algal filament less than the first algal filament is spent, the second algal filament is seeded on supporting layer and forms alga cells layer.Due in algae
In cellular layer, the length of the second algal filament is shorter, therefore in the alga cells layer for being attached to supporting layer, exists between the second algal filament
More gap, these gaps are interconnected, and partial gap can go directly the surface of alga cells layer.Therefore, in alga cells layer
During culture, even if alga cells generate more oxygen or thin to algae during the cultivation process by photosynthesis
Born of the same parents' layer provides the gases such as excessive carbon dioxide, and the gases such as these oxygen and carbon dioxide also can be by above-mentioned gap from algae
It is escaped in class cellular layer, to be not easy to accumulate in alga cells layer, and then the production being bubbled in alga cells layer is alleviated or avoided
It is raw.Therefore, compared with frequently generating bubbling in the prior art during the cultivation process alga cells layer, the present invention can reduce or keep away
Exempting from the bubbling in alga cells layer makes the phenomenon that cells of superficial layer is far from supporting layer, so that cells of superficial layer is easy to from supporting layer
It obtains culture medium and finally significantly improves the cultured output of alga cells so that cells of superficial layer can be grown more vigorously.
In addition, the bottom cell in alga cells layer is covered by cells of superficial layer in the Immobilized culture of the prior art, lead to
Chang Wufa obtains the gases such as sufficient illumination and carbon dioxide, therefore in the incubation of alga cells, bottom cell base
Originally in a dormant state, almost without growth.And in embodiments of the present invention, as shown in the above, in alga cells layer
There are more gap, and partial gap can go directly the surface of alga cells layer, so that extraneous light and carbon dioxide etc.
Gas can be entered inside alga cells layer by these gaps, and bottom cell is enabled to obtain a certain amount of illumination and dioxy
Change the gases such as carbon, and then a degree of growth can be carried out using gases such as the illumination and carbon dioxide.Therefore, the present invention is real
The cultural method of the alga cells of example offer is provided, additionally it is possible to cultured output provided by alga cells layer indsole confluent monolayer cells is improved,
To further increase total cultured output of the cultural method of alga cells.
It is understood that the length of the second algal filament is shorter, gap present in alga cells layer is more, more can
Mitigate the bubbling phenomenon occurred in alga cells layer, but the difficulty for shorten processing to the first algal filament also increases accordingly, therefore,
For above-mentioned technical proposal, the 1/5-1/2 that the length of the second algal filament formed after processing is the length of the first algal filament is shortened,
To which on the basis of mitigating the bubbling phenomenon occurred in alga cells layer well, reduction carries out the first algal filament to shorten processing
Difficulty.For example, 2-3 the second algal filaments can be divided into the first algal filament in shortening processing, the length of these the second algal filaments is big
The about 1/3-1/2 of the length of the first algal filament.It should be noted that the embodiment of the present invention and being not particularly limited the length of the second algal filament
Degree, those skilled in the art can according to the cost for shortening processing and shorten processing can achieve the effect that, setting the second algal filament
Length.
Since in the incubation of alga cells, the alga cells of same level have substantially in alga cells layer
Identical extent of growth, thus, in order to make above-mentioned the second algal filament for being located on the same floor face extent of growth during the whole culture process
It is roughly the same, therefore, in embodiments of the present invention, it can further make the equal length of each second algal filament, so that adhering to
In the cellular layer on supporting layer, the length for being located on the same floor second algal filament in face keeps substantially phase during the whole culture process
Deng so that in following incubations, it is not necessary to due to the length of the second algal filament in alga cells layer is different repeatedly repeatedly
Ground is handled processing such as (i.e. following) cutting out, cut or harvest to the second algal filament of different length, finally makes alga cells
Cultural method be simplified.
It in embodiments of the present invention, include that machinery shortens processing mode for the processing that shortens of the first algal filament, machinery shortens
Processing mode includes: that the first algal filament is cut out, cuts or is stirred, so that the first algal filament is broken, to form the second algal filament.I.e.
Machinery shortens processing and includes cutting-out method, patterning method and paddling process, specifically:
When carrying out shortening processing to the first algal filament using cutting-out method, usually usable scissors cuts the first algal filament
It cuts out, and multiple first algal filaments arranged side by side can be cut out simultaneously.
, can be by knife-edges such as choppers when carrying out shortening processing to the first algal filament using patterning method, disposable cutting is more
A first algal filament, particularly, for it is certain it is intricate, be not easy the first regularly arranged algal filament, special algal filament can be manufactured and cut
Cutter tool, algal filament cutting utensil may include bottom plate, and bottom plate is equipped with the blade of multiple strips, and the blade of these strips is in bottom plate
On be arranged across in length and breadth, form the network that is made of blade, the blade of blade is directed toward the direction for deviating from bottom plate, adjacent
The distance between two blades be less than preset second algal filament minimum length, in this way, need to it is above-mentioned be not easy it is regularly arranged
The first algal filament when being cut, which can be shakeout as much as possible in the plane on such as ground, then will be above-mentioned
Algal filament cuts utensil left-hand thread (makes blade be directed toward the first algal filament) on the first algal filament, and then these first algal filaments are cut into length
The second shorter algal filament；Further for not scissile first algal filament, same position of the sharp object on the first algal filament also can be used
Firmly scribing repeatedly is set, so that the first algal filament disconnects at this location, the second shorter algal filament of formation length.
When carrying out shortening processing to the first algal filament using paddling process, the first algal filament can be placed in biggish container, and
The liquid such as water or culture medium are added, then it is stirred, until the first algal filament therein is fractured into length lesser second
Until algal filament；Dedicated mixing tool can be used to carry out for the above process, for example, rotatable knife can be arranged in large container
Piece rotates blade by devices such as motor, cuts off the first algal filament in container, it should be noted that the embodiment of the present invention
It is preferred that being manually stirred to the first algal filament, preferably to control the length of the second algal filament as made of the fracture of the first algal filament.
It is understood that under the premise of the first algal filament is easy to regularly arranged, it is preferred to use cutting-out method or patterning method pair
First algal filament carries out shortening processing, and carries out cutting out or cutting for rule as far as possible, so that the section of the second algal filament is neat, and disconnected
The area in face is smaller, to reduce the damage of the alga cells to the second algal filament section part as far as possible.
In addition, the processing that shortens for the first algal filament may also include nature and shorten processing mode, processing mode is shortened naturally
It include: to provide culture medium, carbon dioxide gas and illumination to the first algal filament, and it is default to be greater than the ventilation ratio of carbon dioxide gas
The first ventilation ratio, and/or make illumination intensity of illumination be greater than preset first intensity of illumination so that the first algal filament grows and divides
It splits for multiple second algal filaments.First ventilation ratio and the first intensity of illumination cultivate the alga cells quantity in volume by unit and determine,
For example, the first ventilation ratio can be 1vvm, and the first intensity of illumination can be 800 μm of ol/m for most algae cell2/ s,
In specific incubation, the first algal filament can be placed in culture medium, be greater than 1vvm according to ventilation ratio and be passed through dioxy into culture medium
Change carbon gas, or is greater than 800 μm of ol/m according to intensity of illumination2/ s provides illumination to the first algal filament, and above-mentioned ventilation ratio is much larger than algae
Ventilation ratio needed for class cell normal growth, this division for causing alga cells extremely vigorous, but its volume can not increase, from
And promptly to be split into the second shorter algal filament of multiple length in the first algal filament；Similarly, above-mentioned intensity of illumination can also make
It obtains the first algal filament and is promptly split into the second shorter algal filament of multiple length.It, can be by gauze to mixed after the completion of the above process
There are the first algal filament and the culture medium of the second algal filament to be filtered, longer first algal filament and the second shorter algal filament are separated, into
And the second shorter algal filament is collected, and continue to cultivate to longer first algal filament.Above-mentioned culture and filter process is repeated several times,
It can be obtained multiple second algal filaments.
Referring to Fig. 2, the mechanical dissection that machinery shortens in processing may injure in shortening in processing for the first algal filament
The alga cells of segmentation portion in first algal filament, and shortening abnormal vigorous division in processing naturally may also injure in the first algal filament
Cell activity, so that there may be the alga cells that come to harm of activity in the second algal filament.Therefore, in subsequent culture
In the process, the growth of the alga cells in the second algal filament is likely to occur exception.In order to avoid the generation of above-mentioned abnormal phenomenon, at this
In inventive embodiments, after step 20 (carrying out the step of shortening processing to the first algal filament), step 30 (is connect the second algal filament
The step of planting on supporting layer, obtaining the alga cells layer being attached on supporting layer) before, the cultural method of alga cells also wraps
It includes:
Step 21 provides culture medium, carbon dioxide gas and illumination to the second algal filament, and makes the ventilation of carbon dioxide gas
Than be less than preset second ventilation ratio, and/or make illumination intensity of illumination be less than preset second intensity of illumination so that second
The activity for the alga cells being damaged in algal filament because shortening processing is restored, and the process of step 21 is known as renewal cultivation.
Wherein, the definition of ventilation ratio and intensity of illumination is identical as above content, and details are not described herein again, according to alga cells and practical cultivation
The difference of environment has different settings for the second ventilation ratio and the second intensity of illumination, for example, for most of alga cells
For, the second ventilation ratio can be 0.2vvm, and the second intensity of illumination can be 50 μm of ol/m2/ s, since the second intensity of illumination is lower than algae
Intensity of illumination needed for class cell normal growth, thus by above-mentioned setting, so that alga cells not can be carried out growth substantially, only
Self structure can be repaired using carbon dioxide gas and illumination, restore cell activity.It should be noted that in step 21,
When intensity of illumination is 0, alga cells can still be repaired activity itself by respiration, but ventilation ratio is not under normal conditions
It can be 0.
Fig. 3 to Fig. 6 is please referred to, for above-mentioned technical proposal and its improvement project, step 40 is (i.e. to alga cells layer
In alga cells in the second algal filament the step of being cultivated) include:
Step 41 provides culture medium, carbon dioxide gas and illumination to alga cells layer, and keeps the intensity of illumination of illumination big
In the second intensity of illumination, so that the algae cell growth in the second algal filament.Specifically, culture medium may include water, nutriment and
Growth factor, wherein nutriment may include carbon source, nitrogen source, microelement and metal salt etc.；Culture is provided to alga cells layer
The concrete mode of base includes but is not limited to spray, trickle irrigation, infiltration and atomization etc.；The carbon dioxide gas provided to alga cells can
It can also be the mixed gas including carbon dioxide gas, such as air, specific dioxy for pure carbon dioxide gas
Changing carbon amounts can be configured according to the alga cells actually cultivated；Second intensity of illumination is identical as content mentioned above, by algae
Class cell determines, can be 50 μm of ol/m2/ s, in embodiments of the present invention, for most of alga cells, preferred illumination is strong
Degree is 200 μm of ol/m2/s-800μmol/m2/s。
Referring to Fig. 3, the second algal filament growing location in alga cells layer is too long, causes to avoid in long time cultivation
The hidden danger of bubbling is generated in alga cells layer, step 40 (carries out the alga cells in the second algal filament in alga cells layer
The step of culture) further include:
Step 42a, when the length of the second algal filament in alga cells layer is greater than preset length；Default length is greater than to length
Second algal filament of degree is cut out, cuts or harvests, so that too long the second algal filament in the part shortens or to leave algae thin
Born of the same parents' layer.Specifically, for example, by the observation alga cells layer such as naked eyes or microinstrument, when length is greater than in advance in alga cells layer
If the second algal filament of length is more, the second algal filament that the length is greater than preset length is cut off or cut out, it is made to be fractured into length
Shorter the second new algal filament, or the second algal filament that length is greater than preset length is harvested, wherein not according to alga cells
Together, the size of preset length is different, can generally be determined according to the length of the first algal filament.It should be noted that step 42a can
Implemented repeatedly according to the length situation of change of the second algal filament in alga cells layer.By above-mentioned setting, prevent the second algal filament from growing
Too long to length and cause to generate bubbling in alga cells layer, and then the cultured output for avoiding bubbling phenomenon from causing alga cells drops
It is low.
Referring to Fig. 4, in addition to longer second algal filament of partial-length is handled in alga cells layer, it can also be by algae
All second algal filaments in class cellular layer all harvest, and specifically, step 40 is (i.e. to the algae in the second algal filament in alga cells layer
The step of class cell is cultivated) further include:
Step 43a, when the length of the second algal filament in alga cells layer is greater than preset length, in alga cells layer
All the second algal filament is harvested；Specific collecting method is described in detail in the above content, details are not described herein again.
Step 45a, whole second algal filaments are mixed uniformly, then all the second algal filaments are seeded on supporting layer.It can be with
Understand, in the alga cells culture of step 41, the cells of superficial layer in alga cells layer can obtain sufficient illumination and
Carbon dioxide gas, so that growth is more vigorous, then the length of the second algal filament in cells of superficial layer is longer, and bottom cell can not
Enough illumination is obtained, therefore the length of the second algal filament in bottom cell is shorter.Therefore, in step 45a, by that will harvest
Renewed vaccination forms new alga cells layer, so that new alga cells on supporting layer after all second algal filaments afterwards mix
The ratio of longer second algal filament reduces in cells of superficial layer in layer, so that reducing longer second algal filament causes new algae
A possibility that bubbling is generated in cellular layer, thereby reduces the risk for being bubbled and impacting to the cultured output of alga cells.
Step 46a, the alga cells in the second algal filament being seeded on supporting layer are cultivated.It is specific in the step
Cultural method with it is identical to the cultural method of alga cells in step 41, details are not described herein again.
Scheme is advanced optimized as what above-mentioned all second algal filaments by alga cells layer all harvested, in step
After 43a, before step 45a, step 40 (the step of cultivating the alga cells in the second algal filament in alga cells layer) is also
Can include:
Step 44a, the second algal filament for being greater than preset length to length in all the second algal filaments carries out shortening processing.By this
Step will harvest longer second algal filament of length in obtained all the second algal filaments and handle second algae shorter for new length
Silk, thus in subsequent step 45 so that the length of the second algal filament in the alga cells layer being seeded on supporting layer compared with
It is short, it further prevents generating more bubbling because the second algal filament growing location is too long in alga cells layer, finally avoids bubbling pair
The cultured output of alga cells has an impact.
Referring to Fig. 5, it is understood that in most cases, the embodiment of the present invention can be avoided alga cells layer
The generation of middle bubbling, but in the few cases (such as too fast situation or alga cells of the speed of growth of certain alga cells
The too long situation of incubation time) under, there are the possibility that bubbling is generated in alga cells layer.In order to these issuable bubblings
It is handled, in embodiments of the present invention, step 40 (cultivates the alga cells in the second algal filament in alga cells layer
The step of) further include:
Step 42b, it when generating bubbling in alga cells layer, aligns the second algal filament stated at bubbling and is cut out, cuts
Or harvesting, it disappears so as to be bubbled.Specifically, for example, working as alga cells by the observation alga cells layer such as naked eyes or microinstrument
When there are multiple bubblings in layer, the second algal filament at the bubbling is cut off or cut out, it is made to be fractured into shorter the second new algae of length
Silk, or the second algal filament at the bubbling is harvested.It should be noted that step 42b can be roused according in alga cells layer
The case where bubble, implements repeatedly.By above-mentioned setting, the bubbling is handled in time when generating bubbling in alga cells layer, is kept away
Exempting from bubbling phenomenon causes the cultured output of alga cells to reduce.
Referring to Fig. 6, in addition to generating the second algal filament at bubbling to alga cells layer and handling, it can also be thin by algae
All second algal filaments in born of the same parents' layer all harvest, and specifically, step 40 is (i.e. thin to the algae in the second algal filament in alga cells layer
The step of born of the same parents cultivate) further include:
Step 43b, when generating bubbling in alga cells layer, all the second algal filaments in alga cells layer are adopted
It receives；Specific collecting method has been described in detail in the above content, and details are not described herein again.
Step 45b, whole second algal filaments are mixed uniformly, then all the second algal filaments are seeded on supporting layer.It can be with
Understand, in the alga cells culture of step 41, the cells of superficial layer in alga cells layer can obtain sufficient illumination and
Carbon dioxide gas, so that growth is more vigorous, then the length of the second algal filament in cells of superficial layer is longer, and bottom cell can not
Enough illumination is obtained, therefore the length of the second algal filament in bottom cell is shorter.Therefore, in step 45b, by that will harvest
Renewed vaccination forms new alga cells layer, so that new alga cells on supporting layer after all second algal filaments afterwards mix
The ratio of longer second algal filament reduces in cells of superficial layer in layer, so that reducing longer second algal filament causes new algae
A possibility that bubbling is generated in cellular layer, thereby reduces the risk for being bubbled and impacting to the cultured output of alga cells.
Step 46b, the alga cells in the second algal filament being seeded on supporting layer are cultivated.It is specific in the step
Cultural method is identical with the method cultivated in step 41 to alga cells, and details are not described herein again.
Further, after step 43b, before step 45b, step 40 is (i.e. to the algae in the second algal filament in alga cells layer
The step of class cell is cultivated) further include:
Step 44b, the second algal filament for being greater than preset length to length in all the second algal filaments carries out shortening processing.By this
Step will harvest longer second algal filament of length in obtained all the second algal filaments and handle second algae shorter for new length
Silk, thus in subsequent step 45 so that the length of the second algal filament in the alga cells layer being seeded on supporting layer compared with
It is short, it further prevents generating more bubbling because the second algal filament growing location is too long in alga cells layer, finally avoids bubbling pair
The cultured output of alga cells has an impact.Wherein, preset length depends on the speed of growth and alga cells layer of alga cells
Concentration, it is however generally that can be set to the average length of the first algal filament.
In step 30 (the second algal filament is seeded on supporting layer, obtains the alga cells layer being attached on supporting layer
Step) before, the cultural method of alga cells further includes building immobilization cultivation platform, this builds the specific of immobilization cultivation platform
Process are as follows:
Step 11 builds supporting layer.Supporting layer can be selected flexible material (such as silk), water wetted material (such as timber) or
Flexible water wetted material (such as cotton), flexible material can adapt to the culture environment of various kinds, and water wetted material has good guarantor
Hold the performance of culture medium, the advantages of both flexible water wetted material then has both, according to the culture demand of actual alga cells, ability
Field technique personnel can suitably select suitable supporting layer.
Culture medium storage container is arranged in step 12, stores culture medium, culture medium storage container in culture medium storage container
The large-scale sealing liquid storage container such as abacterial water tank can be selected, to ensure the Preservation in sterile condition of culture medium.
Culture medium conveying device is arranged in step 13 between supporting layer and culture medium storage container, so that culture medium stores
Container provides culture medium to supporting layer, and supporting layer is made to return to the culture medium not being consumed to culture medium storage container.Culture medium
Conveying device can be made of a plurality of pipeline, circulating pump, and specific setting can cultivate working environment and the training of platform according to immobilization
The actual parameter for supporting base circular flow determines that the present invention is particularly limited not to this.
It is understood that step 11 to step 13 can carry out before step 20, it can also be after step 20, before step 30
It carries out, can additionally be carried out simultaneously with step 20.This is because building immobilization cultivation platform and the first algal filament shortens processing
It does not interfere with each other, i.e., before the inoculation of the second algal filament, step 11 can be carried out at any time to step 13.The embodiment of the present invention preferably walks
Rapid 11 carry out before step 20 to step 13, with ensure by shorten processing the second algal filament is made in the first algal filament after, and
When the second algal filament is seeded on supporting layer.
For above-mentioned technical proposal and its improvement project, in step 40 (i.e. in the second algal filament in alga cells layer
Alga cells the step of being cultivated) after, the cultural method of alga cells further include:
Step 50 harvests the second algal filament in alga cells layer.Above content be already mentioned above to the second algal filament into
The method of row harvesting, the harvesting of the second algal filament is carried out using the above method, thin for polysaccharide, protein and triglycerides etc.
The collecting method of intracrine object, same as the prior art, details are not described herein again.For the second algae of length obtained by harvesting
Silk, can directly be used as the first algal filament, the training of alga cells provided in an embodiment of the present invention is again carried out since step 20
The method of supporting improves the culture efficiency of alga cells to realize the Cyclic culture of alga cells.
It is thin that algae is improved possessed by the cultural method for the alga cells that embodiment provides in order to further illustrate the present invention
The effect of the cultured output of born of the same parents, present inventor have also carried out following comparative experiments:
Comparative experiments 1:
In the comparative experiments, experimental group using raw material Huang silk algae as the first algal filament, control group using raw material Huang silk algae as
Raw material algal filament, using filter paper as supporting layer.
Step 10, tiling is identical as poly (methyl methacrylate) plate size on the poly (methyl methacrylate) plate that length × width is 1m × 1m, thickness is 5cm
Non-woven fabrics, and identical with the non-woven fabrics size filter paper of tiling on non-woven fabrics, then by the poly (methyl methacrylate) plate together with thereon
Non-woven fabrics and filter paper place (while taking measures to prevent non-woven fabrics and filter paper from falling off) vertically, non-woven fabrics and poly (methyl methacrylate) plate it
Between, culture medium is constantly added dropwise from the two top, culture medium leaves from the other end, falls into the sink below poly (methyl methacrylate) plate
Interior, which is culture medium storage container, and immersible pump and pipeloop is arranged, and constitutes culture medium conveying device, so that culture
Base can carry out circulation infiltration to filter paper (a set of above-mentioned apparatus is respectively arranged in experimental group and control group).
Step 20 filters the yellow silk algae obtained by Liquid Culture, remove liquid, obtain Huang Sizao algal gel or
Algae piece, i.e. raw material Huang Sizao；Then:
(1) it experimental group: using the raw material Huang silk algae as the first algal filament, with scissors by the first algal filament elder generation lateral shear, then indulges
To shearing, the first algal filament rule as far as possible is made to shorten, to form the second algal filament；
(2) control group: using the raw material Huang silk algae as raw material algal filament.
Step 30 dissolves the raw material algal filament of the second algal filament of experimental group and control group in the medium respectively, then will
Culture medium dissolved with the second algal filament and the culture medium dissolved with raw material algal filament are sprayed at respectively on the filter paper of different supporting layers,
Complete the inoculation of the second algal filament and raw material algal filament.
It is passed through the mixed gas of 2% carbon dioxide and air in step 40, the culture medium into sink, and passes through aeration stone
The mixed gas is dispersed, the dissolution of the mixed gas is promoted；The circulation of culture medium is carried out using culture medium conveying device,
So that filter paper and alga cells layer thereon are persistently cultured base and are infiltrated；Meanwhile using fluorescent lamp to being attached on filter paper
The continuous surface of alga cells layer is irradiated.
Step 50, after persistently cultivating three days to the second algal filament by step 40, respectively by the filter of experimental group and control group
Alga cells on paper are all swept away with water, are mixed, and the salinity and impurity mixed in alga cells is cleaned, and then pass through suction filtration side
Method obtains algal gel, and algal gel is put into 105 DEG C of baking ovens and is dried, then weighs respectively and calculates yield.
Table 1
| Incubation time | Culture area | First algal filament/raw material algal filament length | Average product | |
| Experimental group | 4 days | 1m2 | It is short | 20g/m2/ day |
| Control group | 4 days | 1m2 | It is long | 17g/m2/ day |
Experimental result
In comparative experiments 1, the first algal filament of the Huang Sizao of experimental group by shortening processing, obtain length it is shorter second
Then second algal filament is seeded on filter paper by algal filament；The raw material algal filament of the Huang Sizao of control group is simultaneously untreated, is directly inoculated with
On filter paper.From the point of view of experimental phenomena, it is nearly free from bubbling in the alga cells layer of experimental group, and the alga cells of control group
The bubbling generated in layer is more；It can be obtained according to table 1, the average product of experimental group is apparently higher than the average product of control group, thus
Prove that the cultural method of alga cells provided in an embodiment of the present invention can actually improve the cultured output of alga cells.
Comparative experiments 2:
In the comparative experiments, experimental group using raw material spirulina as the first algal filament, control group using raw material spirulina as
Raw material algal filament, using glass fibre membrane as supporting layer.
Step 10, tiling and poly (methyl methacrylate) plate size on the poly (methyl methacrylate) plate that length × width is 0.3m × 0.3m, thickness is 5cm
Identical non-woven fabrics, then place the poly (methyl methacrylate) plate vertically together with non-woven fabrics thereon (while taking measures to prevent nonwoven
Cloth falls off), culture medium is constantly added dropwise between non-woven fabrics and poly (methyl methacrylate) plate, from the two top, culture medium leaves from the other end,
It falls into the sink being located at below poly (methyl methacrylate) plate, which is culture medium storage container, immersible pump and pipeloop are set,
Culture medium conveying device is constituted, enables culture medium to recycle non-woven fabrics and filter paper in infiltration supporting layer and (experimental group and compares
Each a set of above-mentioned apparatus of setting of group).
Step 20,
(1) experimental group: Liquid Culture is carried out to the first algal filament, and excessive illumination and excess carbon dioxide gas are provided, is obtained
The second algal filament is obtained, the spiral number on the second algal filament is 3-4；
(2) control group: not handled raw material algal filament, and the spiral number on raw material algal filament is 6-8, and length is compared with the second algae
Silk is longer.
Step 30, the inoculation method by suction filtration connect the raw material algal filament of the second algal filament of experimental group and control group respectively
On kind to the circular glass tunica fibrosa of diameter 45mm, four circular glass tunica fibrosas for being inoculated with the second algal filament are arranged in experimental group,
Four glass fibre membranes for being inoculated with raw material algal filament are arranged in control group, are then respectively placed in the immobilization cultivation platform put up
On non-woven fabrics.
It is passed through the mixed gas of 2% carbon dioxide and air in step 40, the culture medium into sink, and passes through aeration stone
The mixed gas is dispersed, the dissolution of the mixed gas is promoted；The circulation of culture medium is carried out using culture medium conveying device,
So that glass fibre membrane and alga cells layer thereon are persistently cultured base and are infiltrated in experimental group and control group；Meanwhile it using
Fluorescent lamp carries out prolonged exposure to the surface for the alga cells layer being attached on glass fibre membrane.
(1) experimental group: after 2 days, the respective location of the alga cells layer on 4 glass fibre membranes is bubbled, and passes through use
The picking methods that water rinses, the second algal filament on each glass fibre membrane of experimental group is harvested respectively, and the inoculation by filtering
The second algal filament that harvesting obtains is seeded on each glass fibre membrane by method again；
(2) control group: after 2 days, multiple positions of the alga cells layer on 4 glass fibre membranes are bubbled, not to this
Bubbling is handled.
Above-mentioned condition of culture is still used, respectively the raw material algal filament of the second algal filament of experimental group and control group is continued to train
It supports.
Step 50, Immobilized culture are total carry out 4 days after, respectively by the alga cells on the glass fibre membrane of experimental group,
It is swept away with the alga cells on the glass fibre membrane of control group with water, washes away the salinity and impurity of frustule remaining, then carry out
Drying and weighing, calculate yield.
Table 2
| Incubation time | Culture area | First algal filament/raw material trichome length | Average product | |
| Experimental group | 4 days | 0.006m2 | First algal filament is shorter, 3-4 spiral | 15g/m2/ day |
| Control group | 4 days | 0.006m2 | Raw material algal filament is longer, 6-8 spiral | 12g/m2/ day |
Experimental result
In comparative experiments 2, the length of the second algal filament of experimental group is more shorter than the length of the raw material algal filament in control group；And
And after Immobilized culture carries out 2 days, the second algal filament on glass fibre membrane is harvested and is re-started inoculation by experimental group, is made
It obtains in the alga cells layer for being attached to glass fibre membrane, the length being made of the preferable cells of superficial layer of growth conditions is increased
Second algal filament and constant the second algal filament of the length being made of the poor bottom cell of growth conditions re-mix, to reduce
The ratio of longer second algal filament in the surface layer of alga cells layer, to reduce issuable bubbling in follow-up cultivation.From reality
It tests from the point of view of phenomenon, alga cells layer only produces less bubbling in experimental group, and alga cells layer is only in culture 2 in control group
A large amount of bubbling is just produced after it, and bubbling phenomenon becomes even more serious after culture 4 days；Can be obtained according to table 2, experimental group compared with
Average product it is higher than the average product of control group by 25%, illustrate generation that control is bubbled to the cell cultured output of spirulina
Improving has apparent facilitation.
All the embodiments in this specification are described in a progressive manner, the same or similar between each embodiment
Part may refer to each other, and each embodiment focuses on the differences from other embodiments.
The above description is merely a specific embodiment, but scope of protection of the present invention is not limited thereto, any
Those familiar with the art in the technical scope disclosed by the present invention, can easily think of the change or the replacement, and should all contain
Lid is within protection scope of the present invention.Therefore, protection scope of the present invention should be based on the protection scope of the described claims.
Claims (9)
1. a kind of cultural method of alga cells, which is characterized in that the cultural method of the alga cells includes:
Shorten processing to the first algal filament, obtain the second algal filament that length is less than first algal filament, first algal filament and
Second algal filament is made of alga cells；
Second algal filament is seeded on supporting layer, the alga cells layer being attached on the supporting layer is obtained；
Alga cells in second algal filament described in the alga cells layer are cultivated；
Described to shorten processing be that nature shortens processing, described to shorten processing naturally and include:
Culture medium, carbon dioxide gas and illumination are provided to first algal filament, and makes the ventilation ratio of the carbon dioxide gas
Greater than preset first ventilation ratio, and/or the intensity of illumination of the illumination is made to be greater than preset first intensity of illumination, so that described
First algal filament grows and is split into multiple second algal filaments；
Preset first ventilation ratio is ventilation ratio needed for alga cells normal growth；Preset first intensity of illumination
For intensity of illumination needed for alga cells normal growth；
The alga cells in the second algal filament described in the alga cells layer carry out culture and specifically include:
Culture medium, carbon dioxide gas and illumination are provided to the alga cells layer, and is greater than the intensity of illumination of the illumination
Second intensity of illumination, so that the algae cell growth in second algal filament；
Intensity of illumination needed for second intensity of illumination is lower than alga cells normal growth；
When generating bubbling in the alga cells layer, second algal filament being located at the bubbling is cut out, is cut
Or harvesting, so that the bubbling disappears.
2. the cultural method of alga cells according to claim 1, which is characterized in that the length of second algal filament is institute
State the 1/5-1/2 of the length of the first algal filament.
3. the cultural method of alga cells according to claim 1, which is characterized in that it is described to first algal filament into
It is described second algal filament is seeded in the supporting layer to go forward after row shortens processing, further includes:
Culture medium, carbon dioxide gas and illumination are provided to second algal filament, and makes the ventilation ratio of the carbon dioxide gas
Less than preset second ventilation ratio, and/or the intensity of illumination of the illumination is made to be less than preset second intensity of illumination, so that in institute
It states in the second algal filament because the activity of alga cells being damaged due to processing that shortens is restored；
Ventilation ratio needed for second ventilation ratio is lower than alga cells normal growth；
Intensity of illumination needed for second intensity of illumination is lower than alga cells normal growth.
4. the cultural method of alga cells according to claim 1, which is characterized in that described in the alga cells layer
Alga cells in second algal filament are cultivated further include:
When the length of second algal filament in the alga cells layer is greater than the average length of the first algal filament, length is greater than
Second algal filament of the preset length is cut out, cuts or harvests.
5. the cultural method of alga cells according to claim 1, which is characterized in that described in the alga cells layer
Alga cells in second algal filament are cultivated further include:
When the length of second algal filament in the alga cells layer is greater than the average length of the first algal filament, to the algae
Whole second algal filament in cellular layer is harvested；
It mixes whole second algal filaments uniformly, then all second algal filaments is seeded on the supporting layer；
The alga cells being seeded in second algal filament on the supporting layer are cultivated.
6. the cultural method of alga cells according to claim 5, which is characterized in that described to the alga cells layer
In whole second algal filament harvested after, it is described to make before all second algal filaments uniformly mix, it is described to the algae
Alga cells in second algal filament described in class cellular layer are cultivated further include:
It is shortened described in second algal filament progress for being greater than the average length of the first algal filament to length in all second algal filaments
Processing.
7. the cultural method of alga cells according to claim 1, which is characterized in that described in the alga cells layer
Alga cells in second algal filament are cultivated further include:
When generating bubbling in the alga cells layer, whole second algal filament in the alga cells layer is adopted
It receives；
It mixes whole second algal filaments uniformly, then all second algal filaments is seeded on the supporting layer；
The alga cells being seeded in second algal filament on the supporting layer are cultivated.
8. the cultural method of alga cells according to claim 7, which is characterized in that described to the alga cells layer
In whole second algal filament harvested after, it is described to make before all second algal filaments uniformly mix, it is described to the algae
Alga cells in second algal filament described in class cellular layer are cultivated further include:
It is shortened described in second algal filament progress for being greater than the average length of the first algal filament to length in all second algal filaments
Processing.
9. the cultural method of -2 described in any item alga cells according to claim 1, which is characterized in that described by described
Two algal filaments are seeded in the supporting layer and go forward, the cultural method of the alga cells further include:
Build the supporting layer；
Culture medium storage container is set, stores the culture medium in the culture medium storage container；
Culture medium conveying device is set between the supporting layer and the culture medium storage container, so that the culture medium stores
Container provides the culture medium to the supporting layer, and is consumed the supporting layer not to culture medium storage container return
The culture medium.
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