CN104195045A - Method for culturing optical biological cells - Google Patents
Method for culturing optical biological cells Download PDFInfo
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- CN104195045A CN104195045A CN201410401933.1A CN201410401933A CN104195045A CN 104195045 A CN104195045 A CN 104195045A CN 201410401933 A CN201410401933 A CN 201410401933A CN 104195045 A CN104195045 A CN 104195045A
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Abstract
The invention provides a method for culturing optical biological cells, belongs to the field of cell culture, and aims at increasing the yield of optical biological cells. The method for culturing optical biological cells comprises the following steps: inoculating the optical biological cells on a medium by using a first nutrient liquid till the optical biological cells adhered to the medium are not increased; performing light supply culture on the optical biological cells adhered to the medium, providing a second nutrient liquid to the optical biological cells adhered to the medium, and conducting the second nutrient liquid along the direction from the surface layer to the bottom layer of the cells, thereby obtaining cell aggregate of the optical biological cells, wherein the cell aggregate comprises bottom-layer cells which are directly adhered to the medium and cells which are not directly adhered to the medium and are easy to drop off. The method can be applied to production of optical biological cells and/or metabolite of optical biological cells.
Description
Technical field
The present invention relates to field of cell culture, relate in particular to a kind of cultural method of photo bio cell.
Background technology
Extensive, the high yielding culture of photo bio cell are the bottlenecks of its industrialized development of restriction, and exploitation low-cost, high-efficient culture mode is the key of large-scale production.At present, the cultural method of photo bio cell is that water body is cultivated, and is specially in raceway pond or reactor photo bio cell and nutrition, water are mixed, and provide suitable illumination to it, makes its self-sow.But find in actual applications, while adopting water body training method to cultivate photo bio cell, be easy to be subject to the restriction of light utilization efficiency, make it can not meet the demand of current photo bio cell culture technology.
In order to improve better the light utilization efficiency of photo bio cell, method is applied to the cultivation of photo bio cell " to adhere to cultivation ", be specially: photo bio cell attachment is grown in a side of medium, and provide nutritive medium to the opposite side of medium, so that nutritive medium is successively via water distribution layer, medium, provide nutritive medium by cell bottom to top layer again.But in utilizing the process of aforesaid method cultivation photo bio cell, because the character of water distribution layer and dielectric material, cellular layer thickness etc. all can impact the transmission of nutritive medium, make nutritive medium in the process of transmitting, photo bio cell cannot obtain the nutritive medium aequum that growth conditions actual with it matches, and causes the overall yield of photo bio cell not high.
In prior art, the inoculation of photo bio cell many by smearing, the mode such as suction filtration, rely on mechanical force to make it be piled up in dielectric surface, on microcosmic not with medium-tight combination, still there is space in iuntercellular.And the inoculation method that the embodiment of the present invention adopts does not need to photo bio cell and medium to apply mechanical force, but utilize cell self-characteristic (shape, structure, secretory product, surface band is electrical etc.) it is attached on medium spontaneously, more securely (as filiform cell embeds or is hooked in the hole or crinosity structure of medium with being easy to cell amphitrichous; The cell of secretion viscous substance is easy to be combined with medium by being wrapped in the viscous substance of outside, surface is easy to be adsorbed in dielectric surface by electrostatic attraction with the cell of electric charge), like this, the nutritive medium that contains photo bio cell is when dielectric surface constantly flows, to being attached to the continuous shower of cell on medium, make to adhere to firm not cell detachment medium, and the trend of adhering to firm cell cell detachment medium when resisting shower can be combined with medium more closely, firm.
The preferred embodiment of the present embodiment is: in the second nutritive medium, successively add photo bio cell, medium is carried out to repeatedly drip washing, so that photo bio cell attachment is on described medium.Be specially: first photo bio cell is divided into some parts, in the second nutritive medium, add a copy of it, then medium is carried out to cycling elution, until the photo bio cell adding in the second nutritive medium is close to inoculation completely, in the second nutritive medium, add a photo bio cell again, repeat above-mentioned steps until the photo bio cell adhering on described medium after drip washing no longer increases.Adopt in this way inoculation and can judge better that photo bio cell inoculates criticality completely on medium, also can effectively save the consumption of photo bio cell.
In another embodiment of the present invention, photo bio cell is joined in described the second nutritive medium, obtain described the first nutritive medium; Described medium is infiltrated in described the first nutritive medium, make photo bio cell attachment on described medium; Repeat above-mentioned steps, until the photo bio cell adhering on described medium no longer increases.
In this embodiment, make photo bio cell utilize cell self-characteristic (surface band is electrical etc. for shape, structure, secretory product) to be attached on medium in spontaneous mode, to avoid photo bio cell on medium to come off because of flowing of nutritive medium.
The preferred embodiment of the present embodiment is: in the second nutritive medium, successively add photo bio cell, so that medium is repeatedly infiltrated, so that photo bio cell attachment is on described medium.Be specially: first photo bio cell is divided into some parts, in the second nutritive medium, add a copy of it, then medium is infiltrated in described the second nutritive medium that adds photo bio cell, until the photo bio cell adding in the second nutritive medium is close to inoculation completely, in the second nutritive medium, add a photo bio cell again, repeat above-mentioned steps, until the photo bio cell on described medium no longer increases after infiltrating.Adopt in this way inoculation and can judge better that photo bio cell inoculates criticality completely on medium.
It should be noted that, the inoculation method that the embodiment of the present invention adopts is not limited to above-mentioned situation, and those skilled in the art can select most suitable inoculation method according to producing actual needs; In addition, judge that the method that the photo bio cell that adheres on described medium no longer increases can be to observe to be attached with the dielectric surface color of photo bio cell or to detect the photo bio cell content in the second nutritive medium, when being attached with, the dielectric surface color of photo bio cell is identical with photo bio cell color, or the photo bio cell content in the second nutritive medium no longer reduces and shows that the photo bio cell adhering on described medium no longer increases.
In another embodiment of the present invention, in described seeded process, after no longer increasing, the photo bio cell adhering on described medium also comprises: to being attached with the described medium of photo bio cell, carry out to light preculture, obtain being attached to the cell aggregation on described medium; After finishing to light preculture, collect and in described cell aggregation, be not attached directly to the easy disengaging cell on medium; Utilize described the second nutritive medium described medium is carried out to drip washing or described medium is infiltrated in described the second nutritive medium, collect described easy disengaging cell residual on described medium, obtain the described medium that is firmly attached with photo bio cell.
In this embodiment, through above-mentioned seeded process, be attached with photo bio cell on described medium, these photo bio cells utilize self-characteristic to be attached on medium comparatively securely.To these photo bio cells carry out 1-7 days give light preculture, make it be grown to cell aggregation, this cell aggregation comprises the bottom cell being attached directly on medium and is not attached directly to the easy disengaging cell on medium.Here, preferably adopt the mode of scraping to collect described easy disengaging cell.The easy disengaging cell of the mode that adopts scraping on not only can collecting cell aggregate, can also regulate the planeness of the cell aggregation of dielectric surface, thereby obtain evenly good Growth of Cells face.
Further, conventionally also can utilize described the second nutritive medium described medium is carried out to drip washing or medium is infiltrated in the second nutritive medium, to collect easy disengaging cell residual on described medium, be beneficial to and in follow-up cultivation, accurately weigh cell yield.Meanwhile, the trend that does not depart from cell cell detachment medium when resisting shower on medium, can utilize unique characteristics to be combined more closely with medium, firmly, thereby can improve the steadiness of photo bio cell to follow-up cultivation.In another preferred embodiment of above-described embodiment, the described easy disengaging cell of collecting is joined in the second nutritive medium again.In this embodiment, the cell that above-mentioned scraping and drip washing mode can be collected again joins and in the second nutritive medium, is mixed with the first nutritive medium, so that new medium is inoculated, can make preferably so easily to depart from cell and obtain recycle, save the consumption of photo bio cell and nutritive medium.
In another embodiment of the present invention, photo bio cell attachment is in the both sides of described medium.Adhere to while cultivating photo bio cell is one-sided, the second nutritive medium is being passed to by cell surface layer after cell bottom, and the second unnecessary nutritive medium may infiltrate into medium opposite side, for bottom cell, will certainly cause like this loss of the second nutritive medium especially.So, in a preferred embodiment of the invention, photo bio cell is seeded in to medium both sides, even if the second nutritive medium permeates, also can be by the photo bio Cell uptake of opposite side, make implicit more second nutritive medium of bottom iuntercellular of both sides, thus the more close and firm that makes the state of bottom cell better, is combined with medium.
Further, the both sides to described medium provide described the second nutritive medium respectively, and described the second nutritive medium is the direction transmission along cell surface layer to bottom respectively from described medium both sides.Photo bio cell bilateral is being adhered in the process of cultivation, needing the while to provide nutritive medium to the both sides of medium, nutritive medium is being transmitted, so that the photo bio cell of medium both sides all obtains sufficient nutrient liquid from medium both sides along cell surface layer to bottom.Can make so the easy disengaging cell of medium both sides fully grow, and make the bottom cell of medium both sides can obtain the nutritive medium of satisfying the demand.Photo bio cell bilateral is adhered to the output that cultivation can increase substantially photo bio cell and/or its metabolite, can make bottom cell obtain more nutritive medium, being combined better with medium, be more conducive to follow-up culturing process.
In another embodiment of the present invention, described medium is flexible materials, and flexible materials has good snappiness, can adapt to any incubator, to expand the scope of application of the cultural method that the present embodiment provided.
In another embodiment of the present invention, described medium is preferably water wetted material.Select water wetted material can make photo bio cell sooner, better medium is adhered to, thereby raise the efficiency, reduce the loss that may occur in follow-up cultivation.
Further, described medium preferred flexible water wetted material.Flexible water wetted material has flexible and hydrophilic feature concurrently, can either adapt to various incubator, expand the scope of application of the embodiment of the present invention, can make again photo bio cell sooner, combine with medium better, thereby raise the efficiency, reduce the loss that may occur in follow-up cultivation.
Further, described medium is the surperficial described flexible water wetted material with roughness and/or fine hair and/or hole and/or through hole.Because " adhering to cultivation " need improve photo bio cell and/or its metabolite output by expanding bond area.Simultaneously, in order to allow photo bio cell can utilize the self-characteristics such as shape, structure to be attached on medium, be required to be it relatively coarse surface tissue is provided, and hole or through hole can expand significantly medium can bond area, high roughness or tool surface villous is more conducive to photo bio cell medium is adhered to, and so just can make the efficiency of the embodiment of the present invention and the output of unit time be further enhanced.It should be noted that, the medium in the embodiment of the present invention can be cotton, napped cotton fabric, but is not limited to above-mentioned material, and those skilled in the art can select voluntarily according to foregoing and practical situation.
In embodiments of the present invention, photo bio cell is selected from one or more in the photo bio cells such as spherical algae, filamentous algae, arc algae, trilateral algae, polygon algae, grid algae, chlorella, Chlorococcum, haematococcus pulvialis, silkweed and filamentous cyanobacteria.
The present invention can be applicable to the photo bio cell that much cannot inoculate in traditional cultural method, as spherical algae.This is because this cell surface is comparatively smooth, in seeded process, can not form good sticking power with medium.And the inoculation method that adopts the embodiment of the present invention to provide, utilize the secretory product of spherical algae can make this spherical algae spontaneously be attached on medium, then after carrying out to light preculture to spherical frustule, by modes such as scraping and drip washing, be collected in and on medium, adhere to firm not spherical frustule, and originally on medium, adhere to the trend that comparatively firm spherical frustule departs from medium when resisting scraping and shower, can be combined with medium more closely, firm.Be understandable that, the scope of the photo bio cell that the embodiment of the present invention is related is comparatively extensive, and those skilled in the art also can be applied in the method in the frustule of other type according to actual needs.It should be noted that, in the cultivation of photo bio cell, the product of final demand, except photo bio cell itself, can also comprise its metabolite, as materials such as polysaccharide, antioxidant, grease, unsaturated fatty acidss.
In another embodiment of the present invention, after the described cell aggregation that obtains photo bio cell, also comprise: in collecting described cell aggregation, be not attached directly to the easy disengaging cell on medium; Repeat above-mentioned to light cultivation and collection process.Because collection process adopts scraping mode more, still can residual a certain amount of bottom cell on medium, this medium can be equal to the complete medium of inoculation, so just can again to it, carry out cultivating to light, to obtain being attached to the cell aggregation on medium, thereby form working cycle.This working cycle can utilize bottom cell residual on medium better, and obtains higher output when reducing operation.
For the cultural method of photo bio cell provided by the invention is described better, with specific embodiment, be elaborated below.
Summary of the invention
The embodiment of the present invention provides a kind of cultural method of photo bio cell, to improve the output of photo bio cell.
For achieving the above object, embodiments of the invention adopt following technical scheme:
A cultural method for photo bio cell, comprising:
Utilize the first nutritive medium on medium, to inoculate photo bio cell, until the photo bio cell adhering on described medium no longer increases;
The photo bio cell adhering on described medium is carried out cultivating to light, and provide the second nutritive medium to the photo bio cell adhering on described medium, the direction transmission of described the second nutritive medium along cell surface layer to bottom, obtains the cell aggregation of photo bio cell;
Described cell aggregation comprises the bottom cell being attached directly on described medium and is not attached directly to the easy disengaging cell on described medium.
Alternatively, described inoculation comprises:
Photo bio cell is joined in described the second nutritive medium, obtain described the first nutritive medium;
Utilize described the first nutritive medium to carry out drip washing to described medium, make photo bio cell attachment on described medium;
Repeat above-mentioned steps, until the photo bio cell adhering on described medium no longer increases.
Alternatively, described inoculation comprises:
Photo bio cell is joined in described the second nutritive medium, obtain described the first nutritive medium;
Described medium is infiltrated in described the first nutritive medium, make photo bio cell attachment on described medium.
Repeat above-mentioned steps, until the photo bio cell adhering on described medium no longer increases.
Preferably, photo bio cell attachment is in the both sides of described medium.
Further, the both sides to described medium provide described the second nutritive medium respectively, and described the second nutritive medium is the direction transmission along cell surface layer to bottom respectively from described medium both sides.
Alternatively, described in, be seeded in after the photo bio cell adhering on described medium no longer increases and also comprise:
To being attached with the described medium of photo bio cell, carry out to light preculture, obtain being attached to the cell aggregation on described medium;
After finishing to light preculture, collect and in described cell aggregation, be not attached directly to the easy disengaging cell on medium;
Utilize described the second nutritive medium described medium is carried out to drip washing or described medium is infiltrated in described the second nutritive medium, collect easy disengaging cell residual on described medium, obtain the described medium that is firmly attached with photo bio cell.
Further, the described easy disengaging cell of collecting is joined in described the second nutritive medium again.
Preferably, described medium is selected from a kind of in flexible materials, water wetted material and flexible water wetted material.
Further, described medium is the surperficial described flexible water wetted material with roughness and/or fine hair and/or hole and/or through hole.
Alternatively, described photo bio cell is selected from one or more in spherical algae, filamentous algae, arc algae, trilateral algae, polygon algae, grid algae, chlorella, Chlorococcum, haematococcus pulvialis, silkweed and filamentous cyanobacteria.
Alternatively, after the described cell aggregation that obtains photo bio cell, also comprise:
Collect and in described cell aggregation, be not attached directly to the easy disengaging cell on medium;
Repeat above-mentioned to light cultivation and collection process.
The invention provides a kind of cultural method of photo bio cell, the method is seeded in photo bio cell on medium, and the direction of the second nutritive medium along cell surface layer to bottom transmitted.Compared with the conventional method, in the present invention, related nutritive medium supply mode can make the most vigorous easy disengaging cell of growth obtain sufficient nutrient liquid, meanwhile, makes the basic bottom cell in dormant state also can obtain necessary nutritive medium.By nutritive medium is carried out to properly distributed, the output of photo bio cell is improved.
Embodiment
Below the technical scheme in the embodiment of the present invention is clearly and completely described, obviously, described embodiment is only the present invention's part embodiment, rather than whole embodiment.Embodiment based in the present invention, those of ordinary skills, not making the every other embodiment obtaining under creative work prerequisite, belong to the scope of protection of the invention.
The cultural method of the photo bio the cell below embodiment of the present invention being provided is described in detail.
The embodiment of the present invention provides a kind of cultural method of photo bio cell, comprising: utilize the first nutritive medium on medium, to inoculate photo bio cell, until the photo bio cell adhering on described medium no longer increases; The photo bio cell adhering on described medium is carried out cultivating to light, and provide the second nutritive medium to the photo bio cell adhering on described medium, the direction transmission of described the second nutritive medium along cell surface layer to bottom, obtains the cell aggregation of photo bio cell.Described cell aggregation comprises the bottom cell being attached directly on described medium and is not attached directly to the easy disengaging cell on described medium.
In photo bio cell being carried out to the process of " adhering to cultivation ", the character of water distribution layer and dielectric material, cellular layer thickness etc. all can impact nutritive medium in the supply to cell surface layer direction at present.If water distribution layer liquid holdup is large, homogeneous, medium layer wetting ability, permeability are good, are beneficial to nutritive medium to the transmission of bottom cell; And along with the growth of photo bio cell, cellular layer thickness continues to increase, and the intercellular space of part photo bio dwindles, permeability between each confluent monolayer cells is lower (as some photo bio cell arrangement consolidation, some photo bio cell in growth to exocytosis viscous substance etc.), this has limited the transmission of nutritive medium from cell bottom to top layer to a great extent, causes to cell surface layer and supplies with nutrition deficiency.
Corresponding to above-mentioned situation, in embodiments of the present invention, the direction of the second nutritive medium along cell surface layer to bottom permeated, so that easily depart from cell when obtaining more nutritive medium, bottom cell obtains and enough to maintain the nutritive medium of its growth, thereby the overall yield of higher photo bio cell is provided.Be understandable that, to the described described medium that is attached with photo bio cell, provide the concrete mode of the second nutritive medium can have spray, drip irrigation, infiltration, atomization etc., the present invention not described in subtend medium provide the concrete mode of the second nutritive medium to do concrete restriction.
In embodiments of the present invention, medium is to have the medium (as porous material or water wetted material) of holding liquid ability, is absorbed fully guaranteeing in process that nutritive medium permeates in the direction along cell surface layer to bottom by cell.
Described the first nutritive medium is the nutritive medium that is added with photo bio cell.Described the second nutritive medium is not for being added with the nutritive medium of photo bio cell.In the present embodiment, the second nutritive medium comprises water and/or nutritive substance and/or somatomedin, and nutritive substance comprises carbon source, nitrogenous source, trace element, metal-salt etc., and the Carbon and nitrogen sources in this nutritive substance can be with gaseous state (as CO
2, NO
2deng) or liquid (referring to the material of carbon source, nitrogenous source to be dissolved in liquid) form cell is supplied with, preferably with liquid form, supply with.It should be noted that, the growth changes in demand of the photo bio cell that in described culturing process, intensity of illumination, nutraceutical component and each component concentration thereof can be cultivated according to reality regulates.
In embodiments of the present invention, cell aggregation is the final product of gained in culturing process, and this cell aggregation comprises bottom cell and easily departs from cell.Wherein, bottom cell is the cell being attached directly on medium, easily departing from cell is the new cell growing in culturing process, because photo bio cell is successively grown, so easily departing from cell is not attached directly on medium, but be attached on other cells, therefore compare with the bottom cell being attached directly on medium, easily depart from cell and comparatively easily collect.
The embodiment of the present invention provides a kind of cultural method of photo bio cell, and the method is seeded in photo bio cell on medium, and the direction of the second nutritive medium along cell surface layer to bottom transmitted.Compared with the conventional method, the related nutritive medium supply mode of the embodiment of the present invention can make the most vigorous easy disengaging cell of growth obtain sufficient nutrient liquid, meanwhile, makes the basic bottom cell in dormant state also can obtain necessary nutritive medium.By nutritive medium is carried out to properly distributed, the output of photo bio cell is improved.
In another embodiment of the present invention, described inoculation comprises: photo bio cell is joined in described the second nutritive medium, obtain described the first nutritive medium; Utilize described the first nutritive medium to carry out drip washing to described medium, make photo bio cell attachment on described medium; Repeat above-mentioned steps, until the photo bio cell adhering on described medium no longer increases.
Embodiment 1
With ball algae, for photo bio cell to be cultivated, take napped cotton fabric as medium, carry out one side and adhere to cultivation.
Napped cotton fabric is wrapped in to the beaded support surface formation spheroplast that diameter is 10 cm, be immersed in the BG11 nutritive medium containing the 3g/L ball frustule of having an appointment, the mixed gas of carbonic acid gas and air will be passed in this nutritive medium, make this nutritive medium pH value maintain 7 left and right, make spheroplast random motion in nutritive medium, after spheroplast surface is covered with ball algae, spheroplast is taken out in this nutritive medium, put into BG11 nutritive medium rinsing 2 times; Again spheroplast is placed in to nutritive medium, half immerses spheroplast in nutritive medium, second half is exposed to outside nutritive medium, by containing volume fraction, be that the mixed gas of 2% carbonic acid gas passes in nutritive medium, maintain nutritive medium pH in 7 left and right, spheroplast is random rotation under the effect of mixed gas, makes ball frustule attached to it intermittently immerse, leave nutritive medium, and in the outdoor environment in spring of the north, cultured continuously is 10; Collect after the ball frustule on spheroplast surface, continue to cultivate 10; Repeat cultivate to collect step 3 time, approximately 5 grams every square metre of mean yield medium every day.
Embodiment 2
With grid algae, for photo bio cell to be cultivated, take cotton as medium, carry out the two-sided cultivation of adhering to.
The limit that to make containing the BG11 nutritive medium of 1g/L grid frustule of having an appointment be the square cotton of 1 meter along the length of side slowly flows down from both sides, two sides through cotton flow to nutritive medium collecting tank, this nutritive medium that contains grid frustule that flows back into collecting tank is delivered to cotton via submersible pump makes it along two sides of cotton, slowly flow down, form circulation, when the grid frustule in this nutritive medium is exhausted substantially, in this nutritive medium, add grid frustule again, concentration to grid frustule in this nutritive medium rises to about 1g/L, while repeating said process to the 5th, grid frustule content in this nutritive medium no longer reduces, now grid frustule is evenly attached to two sides of cotton, the nutritive medium emptying of grid frustule will be contained in collecting tank, add the nutritive medium that does not contain grid frustule, via submersible pump, make nutritive medium flow through and inoculate complete cotton according to said process, after rinsing 30 min, the nutritive medium emptying of a small amount of cell will be contained in collecting tank, and add nutritive medium, via submersible pump, make nutritive medium according to the flow through cotton of above-mentioned drip washing of said process, by containing volume fraction, be that the mixed gas of 2% carbonic acid gas passes in nutritive medium, maintain nutritive medium pH in 7 left and right, in the outdoor environment in autumn of the north, cultured continuously is 7, collect after the grid frustule on cotton surface, continue to cultivate 7, repeat above-mentioned cultivation collection process three times, approximately 10 grams every square metre of mean yield medium every day.
Obviously, above-described embodiment is only for example is clearly described, and the not restriction to embodiment.For those of ordinary skill in the field, can also make other changes in different forms on the basis of the above description.Here exhaustive without also giving all embodiments.And the apparent variation of being extended out thus or the change protection domain in the invention still.
Claims (11)
1. a cultural method for photo bio cell, is characterized in that, comprising:
Utilize the first nutritive medium on medium, to inoculate photo bio cell, until the photo bio cell adhering on described medium no longer increases;
The photo bio cell adhering on described medium is carried out cultivating to light, and provide the second nutritive medium to the photo bio cell adhering on described medium, the direction transmission of described the second nutritive medium along cell surface layer to bottom, obtains the cell aggregation of photo bio cell;
Described cell aggregation comprises the bottom cell being attached directly on described medium and is not attached directly to the easy disengaging cell on described medium.
2. method according to claim 1, is characterized in that, described inoculation comprises:
Photo bio cell is joined in described the second nutritive medium, obtain described the first nutritive medium;
Utilize described the first nutritive medium to carry out drip washing to described medium, make photo bio cell attachment on described medium;
Repeat above-mentioned steps, until the photo bio cell adhering on described medium no longer increases.
3. method according to claim 1, is characterized in that, described inoculation comprises:
Photo bio cell is joined in described the second nutritive medium, obtain described the first nutritive medium;
Described medium is infiltrated in described the first nutritive medium, make photo bio cell attachment on described medium;
Repeat above-mentioned steps, until the photo bio cell adhering on described medium no longer increases.
4. according to the method described in claim 1-3 any one, it is characterized in that, photo bio cell attachment is in the both sides of described medium.
5. method according to claim 4, is characterized in that, the both sides to described medium provide described the second nutritive medium respectively, and described the second nutritive medium is the direction transmission along cell surface layer to bottom respectively from described medium both sides.
6. according to the method described in claim 1-3 any one, it is characterized in that, described in be seeded in after the photo bio cell adhering on described medium no longer increases and also comprise:
To being attached with the described medium of photo bio cell, carry out to light preculture, obtain being attached to the cell aggregation on described medium;
After finishing to light preculture, collect and in described cell aggregation, be not attached directly to the easy disengaging cell on medium;
Utilize described the second nutritive medium described medium is carried out to drip washing or described medium is infiltrated in described the second nutritive medium, collect easy disengaging cell residual on described medium, obtain the described medium that is firmly attached with photo bio cell.
7. method according to claim 6, is characterized in that, the described easy disengaging cell of collecting is joined in described the second nutritive medium again.
8. method according to claim 1, is characterized in that, described medium is selected from a kind of in flexible materials, water wetted material and flexible water wetted material.
9. method according to claim 8, is characterized in that, described medium is the surperficial described flexible water wetted material with roughness and/or fine hair and/or hole and/or through hole.
10. method according to claim 1, it is characterized in that, described photo bio cell is selected from one or more in spherical algae, filamentous algae, arc algae, trilateral algae, polygon algae, grid algae, chlorella, Chlorococcum, haematococcus pulvialis, silkweed and filamentous cyanobacteria.
11. methods according to claim 1, is characterized in that, after the described cell aggregation that obtains photo bio cell, also comprise:
Collect and in described cell aggregation, be not attached directly to the easy disengaging cell on medium;
Repeat above-mentioned to light cultivation and collection process.
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