seaweed cultivation system

Abstract

A culture system is disclosed, comprising a culture substrate configured to promote seaweed holdup formation and seaweed attachment. The culture system may include one or more of a nutrient phase, a binder, a bioactive agent, and a liquid phase. The culture substrate may be patterned. The culture system can specifically retain and effectively maintain specific seaweed species, such as red algae, kelp, and nori.

Description

[0001] Cross-references to related applications

[0002] This application claims priority to provisional application number 63 / 044,285, filed on June 25, 2020, which is incorporated herein by reference in its entirety for all purposes. Technical Field

[0003] This disclosure generally relates to cultivation systems, and more specifically, to seaweed cultivation systems configured to support the formation of anchorages. Background Technology

[0004] The current process of culturing seaweed from spores involves using textured nylon “culture lines” or “seed lines” to which spores weakly attach during a lab-based seeding process. The culture lines, containing weakly attached juvenile seaweed (gametophytes and sporophytes), are then wound onto ropes in the seaweed farm, which are subsequently placed underwater. This process is inherently variable in terms of yield and throughput, largely due to biofouling (i.e., the seed lines becoming contaminated with unwanted seaweed and other biological species). Biofouling severely reduces seaweed growth and yield. Traditionally, effective anti-biofouling materials (smooth, low-friction films) also reduce seaweed growth and yield due to poor adhesion to these substrates. Other factors affecting yield and throughput include the ease with which the seaweed is damaged, such as by water flow, temperature variations, and nutrient availability. Furthermore, improper packaging and handling can lead to the destruction and loss of juvenile seaweed. Current methods for improving the stability of juvenile seaweed on culture lines focus on the surface texture of existing fibers. In fact, the fibrous texture of the culture line is crucial to the success of seaweed cultivation. A substrate is needed that can provide effective attachment and growth of seaweed while also providing effective resistance to biofouling. Summary of the Invention

[0005] Each implementation involves a culture system configured to retain and effectively maintain spores.

[0006] According to one example (“Example 1”), a culture system includes a culture substrate comprising a low-porosity matrix having a porosity of about 10% or less and having a fibrillated submicron surface structure configured to retain algae by immobilization.

[0007] According to a further example of Example 1 (“Example 2”), the fibrillated submicron surface structure is characterized by an average interfibrillary distance of up to 1000 nm and including 1000 nm.

[0008] According to a further example of Example 1 or Example 2 (“Example 3”), the fibrillated submicron surface structure has an average depth of about 1000 nm or less.

[0009] According to a further example (“Example 4”) of any of Examples 1 to 3, the fibrillated submicron surface structure has an average depth of about 5 nm or less.

[0010] According to a further example (“Example 5”) of any of Examples 1 to 4, the thickness of the low porosity matrix is ​​about 25.4 μm (1 mil) to about 762 μm (30 mil).

[0011] According to a further example (“Example 6”) of any of Examples 1 to 5, the thickness of the low-porosity matrix is ​​about 25.4 μm (1 mil) to about 127 μm (5 mil).

[0012] According to a further example (“Example 7”) of any of Examples 1 to 6, the culture medium is configured as a strip, a matrix (substrate / sheet), a woven product, a nonwoven product, a braided product, a knitted product, a fabric, a particulate dispersion, or a combination of two or more of the above.

[0013] According to a further example (“Example 8”) of any of Examples 1 to 7, the culture substrate includes at least one of the following: a backing layer, a carrier layer, a laminate of multiple layers, a composite material, or a combination thereof.

[0014] According to a further example (“Example 9”) of any of Examples 1 to 8, the low-porosity matrix comprises an expanded fluoropolymer.

[0015] According to a further example of Example 9 (“Example 10”), the expanded fluoropolymer is one of the following: expanded fluorinated ethylene propylene (eFEP), porous perfluoroalkoxyalkane (PFA), expanded ethylene tetrafluoroethylene (eETFE), expanded vinylidene fluoride cotetrafluoroethylene or trifluoroethylene polymer (eVDF-co-(TFE or TrFE)) and expanded polytetrafluoroethylene (ePTFE).

[0016] According to a further example (“Example 11”) of any of Examples 1 to 10, the low-porosity matrix is ​​an expanded polytetrafluoroethylene (ePTFE) matrix.

[0017] According to a further example (“Example 12”) of any of Examples 1 to 8, the low-porosity matrix comprises an expanded thermoplastic polymer.

[0018] According to a further example of Example 12 (“Example 13”), the expanded thermoplastic polymer is one of the following: expanded polyester sulfone (ePES), expanded ultra-high molecular weight polyethylene (eUHMWPE), expanded polylactic acid (ePLA), and expanded polyethylene (ePE).

[0019] According to a further example (“Example 14”) of any of Examples 1 to 8, the low-porosity matrix contains an expanded polymer.

[0020] According to a further example of Example 14 (“Example 15”), the expanding polymer is expanded polyurethane (ePU).

[0021] According to a further example (“Example 16”) of any of Examples 1 to 8, the low-porosity matrix is ​​expanded poly(p-xylene) (ePPX).

[0022] According to a further example of Example 11 (“Example 17”), the PTFE matrix has approximately 0.015 g-mm / m 2 A water vapor permeability coefficient of / day or less, and formed by a method comprising the following steps: (a) preparing a biaxially expanded PTFE film; (b) densifying the expanded PTFE film; and (c) stretching the densified expanded PTFE film.

[0023] According to a further example of Example 18 (“Example 18”), in step (c), the densified expanded PTFE film is stretched at a temperature exceeding the crystallization melting temperature of PTFE.

[0024] According to another further example (“Example 19”) of Example 17 or Example 18, the expanded PTFE film is sintered prior to step (b).

[0025] According to a further example (“Example 20”) of any of Examples 17 to 19, a biaxially expanded PTFE film comprises two or more layers of expanded PTFE.

[0026] According to another further example (“Example 21”) of any of Examples 17 to 20, steps (a)-(c) are performed in a sequential manner.

[0027] According to a further example (“Example 22”) of any of Examples 1 to 21, the culture substrate further comprises a high-porosity matrix having a porosity of at least 30% and a node and fibrillary microstructure, characterized in that the average interfibrillary distance is from about 1 μm to 500 μm, or the average pore size is from about 1 μm to about 500 μm.

[0028] According to a further example of Example 22 (“Example 23”), the high-porosity matrix comprises an expanded fluoropolymer.

[0029] According to a further example of Example 23 (“Example 24”), the expanded fluoropolymer is one of the following: expanded fluorinated ethylene propylene (eFEP), porous perfluoroalkoxyalkane (PFA), expanded ethylene tetrafluoroethylene (eETFE), expanded vinylidene fluoride cotetrafluoroethylene or trifluoroethylene polymer (eVDF-co-(TFE or TrFE)) and expanded polytetrafluoroethylene (ePTFE).

[0030] According to a further example (“Example 25”) of any of Examples 22 to 24, the high porosity matrix is ​​an expanded polytetrafluoroethylene (ePTFE) matrix.

[0031] According to a further example of Example 22 (“Example 26”), the high-porosity matrix comprises an expanded thermoplastic polymer.

[0032] According to a further example of Example 26 (“Example 27”), the expanded thermoplastic polymer is one of the following: expanded polyester sulfone (ePES), expanded ultra-high molecular weight polyethylene (eUHMWPE), expanded polylactic acid (ePLA), and expanded polyethylene (ePE).

[0033] According to a further example of Example 22 (“Example 28”), the high porosity matrix contains an expanded polymer.

[0034] According to a further example of Example 28 (“Example 29”), the expanding polymer is expanded polyurethane (ePU).

[0035] According to another further example of Example 22 (“Example 30”), the high-porosity matrix is ​​expanded poly(p-xylene) (ePPX).

[0036] According to a further example (“Example 31”) of any of Examples 22 to 30, the high-porosity matrix is ​​hydrophobic.

[0037] According to a further example (“Example 32”) of any of Examples 22 to 31, the low-porosity matrix and the high-porosity matrix contain the same material.

[0038] According to a further example (“Example 33”) of any of Examples 22 to 32, the culture substrate is a patterned matrix having patterns of a low-porosity matrix and a high-porosity matrix.

[0039] According to a further example of Example 33 (“Example 34”), the patterns of the low-porosity matrix and the high-porosity matrix are organized or selective patterns.

[0040] According to a further example of Example 33 (“Example 35”), the patterns of the low-porosity matrix and the high-porosity matrix are random patterns.

[0041] According to a further example (“Example 36”) of any of Examples 1 to 35, the culture system includes a nutrient phase associated with at least a portion of the culture substrate.

[0042] According to a further example of Example 36 (“Example 37”), the nutrient phase promotes the growth of algae and / or the attachment of algae to the culture medium.

[0043] According to a further example of Example 36 or Example 37 (“Example 38”), at least a portion of the nutrient phase is entrained in the culture medium, entrained on the culture medium, or entrained in and on the culture medium.

[0044] According to a further example (“Example 39”) of any of Examples 36 to 38, the nutrient phase exists as a coating on the surface of the culture substrate.

[0045] According to a further example (“Example 40”) of any of Examples 1 to 39, the culture medium is provided by a plurality of particles in a dispersion that is formulated for deposition onto a backing layer or carrier matrix.

[0046] According to a further example (“Example 41”) of any of Examples 1 to 40, the culture medium is asymmetric, including a protofibrotic submicron surface structure configured to retain algae on only one side.

[0047] According to another example (“Example 42”), a method for culturing seaweed includes contacting a population of seaweed gametophytes and / or sporophytes with a culture substrate of a culture system of any of Examples 1 to 41 until at least a portion of the seaweed gametophytes and / or sporophytes form an attachment to the nanostructures of the culture substrate.

[0048] According to another example of Example 42 (“Example 43”), the method includes positioning the culture system in an open aquatic environment after a portion of the algal gametophyte and / or sporophyte population has formed an attachment to the culture substrate nanostructure. Attached Figure Description

[0049] The accompanying drawings, which are incorporated in and form part of this specification, are used to aid in a further understanding of this disclosure and illustrate embodiments thereof. Together with the specification, they serve to explain the principles of this disclosure.

[0050] Figure 1A These are photographs depicting naturally occurring algae-bedrock interactions. Source: Morrison L, Feely M, Stengel DB, Blamey N, Dockery P, Sherlock A, Timmins (2009) The attachment of seaweed to bedrock: biophysical evidence for a new paradigm of geoecology, Geobiology 7: 477-487.

[0051] Figure 1B It is by Figure 1A A detailed view of the area marked by the dashed box. Source: Morrison L, Feely M, Stengel DB, Blamey N, Dockery P, Sherlock A, Timmins (2009) Algae attachment to bedrock: biophysical evidence for a new paradigm of geoecology, Geobiology 7: 477–487.

[0052] Figure 2A-2D These are scanning electron microscope (SEM) micrographs taken at various magnifications, depicting the nanostructures of low-porosity matrices according to some embodiments. Figure 2A , Figure 2B , Figure 2C and Figure 2D The scales provided are 100μm, 10μm, 5μm and 5μm.

[0053] Figures 3A-3D These are scanning electron microscope (SEM) micrographs taken at various magnifications, depicting the surface structure of the matrix according to some embodiments. Figure 3A , Figure 3B , Figure 3C and Figure 3D The scales provided are 100μm, 10μm, 5μm and 5μm.

[0054] Figure 4 It is a photograph depicting a network of kelp anchors on a low-porosity matrix surface according to some embodiments.

[0055] Figure 5 This describes, according to some implementation methods, kelp in Figure 2A The growth on the membrane depicted in -D (the two samples on the left) and in Figures 3A-3DA set of photographs depicting growth on a membrane (the two samples on the right).

[0056] Figure 6 This describes, according to some implementation methods, seaweed in Figure 2A The growth on the membrane depicted in -D (the two samples on the left) and in Figures 3A-3D A set of photographs depicting growth on a membrane (the two samples on the right).

[0057] Figure 7 This describes, according to some implementation methods, the red-skinned algae in Figure 2A The growth on the membrane depicted in -D (the two samples on the left) and in Figures 3A-3D A set of photographs depicting growth on a membrane (two samples on the right).

[0058] Figure 8 These are scanning electron microscope (SEM) images depicting the microstructure of a high-porosity matrix according to some embodiments.

[0059] Figure 9 It is a SEM micrograph depicting the microstructure shown in Figure 1, but at a higher magnification.

[0060] Figure 10 These are SEM micrographs depicting the microstructure of a high-porosity matrix according to some embodiments.

[0061] Figure 11 It is a SEM micrograph depicting the microstructure shown in Figure 3, but with a higher magnification.

[0062] Figure 12 This is a schematic diagram depicting the microstructure of a high-porosity matrix according to some embodiments.

[0063] Figure 13 yes Figure 9 Micrographs showing, in sketch form, spores with a diameter of 10 μm or 30 μm that are covered by the interfibrillary space according to some embodiments.

[0064] Figure 14A These are cross-sectional SEM images depicting the microstructure of red algae growing inward into a highly porous matrix according to some embodiments.

[0065] Figure 14B It is shown Figure 14A The image depicts an inward-growing cross-section using SEM microscopy, but at a higher magnification.

[0066] Figure 14C These are cross-sectional optical fluorescence micrographs depicting the microstructure of red algae growing inward into a highly porous matrix according to some embodiments.

[0067] Figure 15 To show the surface SEM micrograph of the microstructure of the high-porosity matrix before sowing with sugar kelp spores according to some embodiments (top image), and to show the optical fluorescence micrograph of the high-porosity matrix after sowing and germination with sugar kelp spores (bottom image).

[0068] Figure 16 Two surface SEM micrographs taken at different magnifications are shown, depicting the inward growth of young red algae into the microstructure according to some embodiments.

[0069] Figure 17 These are surface optical fluorescence micrographs showing the microstructure of red algae growing inward into a highly porous matrix according to some embodiments.

[0070] Those skilled in the art will readily understand that the accompanying drawings referenced herein are not necessarily drawn to scale, but may be enlarged to illustrate various aspects of this disclosure, and in this regard, the drawings should not be considered limiting. Detailed Implementation

[0071] Definitions and Terms

[0072] This disclosure is not intended to be read in a restrictive manner. For example, the terms used in this application should be read broadly in the context of their meanings in the art.

[0073] For imprecise terms, the terms "about" and "approximately" are used interchangeably to indicate that the measured value includes the measured value as well as any measured value reasonably close to the measured value. A measured value reasonably close to the measured value is a reasonably small deviation from the measured value, as understood and readily determined by one of ordinary skill in the art. For example, such deviations may be attributed to measurement errors, differences in the calibration of measuring and / or manufacturing equipment, human error in reading and / or setting measurements, fine-tuning to optimize performance and / or structural parameters due to measurement differences related to other components, specific implementation scenarios, imprecise adjustments and / or manipulations of objects by humans or machines, etc. If it is determined that the value of such a reasonably small difference is not readily determined by one of ordinary skill in the art, the terms "about" and "approximately" can be understood as representing plus or minus 10% of the value.

[0074] Certain terms are used herein for convenience only. For example, terms such as “top,” “bottom,” “upper,” “lower,” “left,” “right,” “horizontal,” “vertical,” “upward,” and “downward” describe only the orientation of the structure or part shown in the figure in its mounting position. In reality, the components referenced can be oriented in any direction. Similarly, in this disclosure, when a process or method is shown or described, the method can be performed in any order or simultaneously unless the context clearly indicates that the method depends on certain actions performed first.

[0075] In the coordinate system shown in the accompanying drawings and described in the specification, the "Y" axis corresponds to the vertical direction, the "X" axis corresponds to the horizontal or transverse direction, and the "Z" axis corresponds to the internal / external direction. Detailed Implementation

[0077] This disclosure relates to a culture system comprising a culture medium. The culture medium is used for the preservation, culture, and / or growth of seaweed, as well as related methods and apparatus. In some embodiments, the culture system is operable to grow seaweed in an open aquatic environment.

[0078] The culture system according to this disclosure can be used for spore culture and growth, as well as the transport and deposition of spores and / or gametophytes / sporophytes. In some embodiments, the culture medium described herein can be used as an improved growth substrate for the growth and culture of various algal forms (e.g., spores, gametophytes, sporophytes) to obtain improved yields and production relative to existing culture practices.

[0079] In some embodiments, the culture system includes a culture substrate, which itself comprises a low-porosity matrix having a fibrillated submicron surface structure on at least one surface of the matrix. The fibrillated submicron surface structure of the low-porosity matrix allows algae to attach to the culture substrate via algal anchorage.

[0080] Holdfasts are root-like structures at the base of seaweed that anchor it to substrates such as rocks. The shape and structure of holdfasts vary by species. Substrate type also influences the shape and structure of holdfasts. Unlike roots of terrestrial plants, seaweed holdfasts do not absorb nutrients; they serve only as anchors.

[0081] Figure 1A The interaction zone between Fucus vesiculosus and the granite bedrock is depicted. The cross-section depicts the anchorage (arrow) and shows the algal lateral branches (1), main axis (2), anchorage area (3), and anchorage-bedrock interface (4). Figure 1B Depicting Figure 1A The detailed view of the area within the dashed box describes in detail the three physicochemically active regions containing the fixation interface. Figure 1BThe arrows indicate rock fragments incorporated into and dispersed within the seaweed holdfast tissue.

[0082] As described in this article, it is surprising to find that nanostructures found on certain low-porosity substrates promote and support the formation of fixatives on the substrate surface. Figure 2A-2D These are SEM micrographs depicting nanostructures on the surface of a low-porosity expanded polytetrafluoroethylene (ePTFE) matrix according to some embodiments. Figure 2A-2C The nanostructures on the first side of the low-porosity matrix were depicted at increased magnification. The scale bar presented is 100 μm. Figure 2A ), 10μm ( Figure 2B ) and 5μm ( Figure 2C ). Figure 2D Nanostructures on the second side of a low-porosity matrix were depicted (scale bar provided: 5 μm). At the lowest magnification ( Figure 2A The surface of the low-porosity matrix appears almost smooth. However, the fibrillated submicron surface structure becomes apparent at higher magnifications. Figure 2B-2D As shown in the figure, the fibrillated submicron surface structure is defined by multiple fibrils. The fibrils define the spaces between them. In some embodiments, such as... Figure 2A-2D As shown, the fibrils of the fibrillated submicron surface structure are interconnected at the nodes. In some embodiments, the fibrillated submicron surface structure contains no nodes, or is substantially node-free.

[0083] The fibrils have a defined average interfibril distance, which in some embodiments can be about 1 nm to about 1000 nm, about 1 nm to about 500 nm, about 1 nm to about 200 nm, about 1 nm to about 50 nm, about 1 nm to about 20 nm, about 1 nm to about 10 nm, about 1 nm to about 5 nm, about 5 nm to about 500 nm, about 5 nm to about 200 nm, about 5 nm to about 100 nm, about 5 nm to about 50 nm, about 5 nm to about 20 nm, about 5 nm to about 10 nm, about 10 nm to about 100 nm, about 10 nm to about 500 nm, about 10 nm to about 10 ... 200nm, approximately 10nm to 100nm, approximately 10nm to approximately 75nm, approximately 10nm to approximately 50nm, approximately 10nm to approximately 25nm, approximately 25nm to approximately 200nm, approximately 25nm to approximately 150nm, approximately 25nm to approximately 100nm, approximately 25nm to approximately 50nm, approximately 50nm to approximately 200nm, approximately 50nm to approximately 150nm, approximately 50nm to approximately 100nm, approximately 100nm to approximately 500nm, approximately 100nm to approximately 200nm, approximately 100nm to approximately 150nm, approximately 150nm to approximately 500nm, or approximately 150nm to approximately 200nm. In some embodiments, the average interfibrillary distance of the fibrils can be about 1 nm, about 2 nm, about 3 nm, about 4 nm, about 5 nm, about 10 nm, about 20 nm, about 30 nm, about 40 nm, about 50 nm, about 60 nm, about 70 nm, about 80 nm, about 90 nm, about 100 nm, about 110 nm, about 120 nm, about 130 nm, about 140 nm, about 150 nm, about 160 nm, about 170 nm, about 180 nm, about 190 nm, about 200 nm, about 250 nm, about 300 nm, about 400 nm, about 500 nm, about 600 nm, about 700 nm, about 800 nm, about 900 nm, or about 1000 nm.

[0084] In some embodiments, the fibrillated submicron surface structures have an average depth of about 1000 nm or less. That is, the fibrillated submicron surface structures exist on the surface of a low-porosity matrix, penetrating the low-porosity matrix to a depth of about 1000 nm or less in the z-axis. In some embodiments, the average depth of the fibrillated submicron surface structures can be about 1 nm to about 1000 nm, about 1 nm to about 500 nm, about 1 nm to about 300 nm, about 1 nm to about 100 nm, about 1 nm to about 50 nm, about 1 nm to about 20 nm, about 1 nm to about 10 nm, about 1 nm to about 5 nm, about 5 nm to about 1000 nm, about 5 nm to about 500 nm, about 5 nm to about 300 nm, about 5 nm to about 100 nm, about 5 nm to about 50 nm, about 5 nm to about 20 nm, about 5 nm to about 10 nm, and about 10 nm to about 1000 nm. 0nm, approximately 10nm to approximately 500nm, approximately 10nm to approximately 300nm, approximately 10nm to approximately 100nm, approximately 10nm to approximately 75nm, approximately 10nm to approximately 50nm, approximately 10nm to approximately 25nm, approximately 25nm to approximately 1000nm, approximately 25nm to approximately 500nm, approximately 25nm to approximately 300nm, approximately 25nm to approximately 100nm, approximately 25nm to approximately 75nm, approximately 25nm to approximately 50nm, approximately 50nm to approximately 1000nm, approximately 50nm to approximately 500nm, approximately 50nm to approximately 300nm, or approximately 50nm to approximately 100nm. In some embodiments, the average depth of the fibrillated submicron surface structure can be about 1 nm, about 2 nm, about 3 nm, about 4 nm, about 5 nm, about 10 nm, about 20 nm, about 30 nm, about 40 nm, about 50 nm, about 60 nm, about 70 nm, about 80 nm, about 90 nm, or about 100 nm.

[0085] In some implementations, the fibrillated submicron surface structure comprises two or more layers of nodes and fibrils. For example, in Figure 2C The text describes a configuration in which the fibrils appear to overlap with other fibrils. In some embodiments, the depth of the fibrillated submicron surface structure represents the layered node and fibril structure.

[0086] In some embodiments, the low-porosity matrix has a porosity of about 10% or less. The porosity of the low-porosity matrix can be about 1% to about 10%, about 1% to about 5%, or about 5% to about 10%. In some embodiments, the porosity of the low-porosity matrix is ​​about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10%.

[0087] The low-porosity matrix may have a thickness of about 1 mil (0.001 inch; 25.4 μm) to about 30 mils (0.03 inch; 254 μm). In some embodiments, the thickness of the low-porosity matrix may be about 1 mil (0.001 inch; 25.4 μm) to about 10 mils (0.01 inch; 254 μm), or about 1 mil (0.001 inch; 25.4 μm) to about 5 mils (0.005 inch; 254 μm). In some embodiments, the thickness of the low-porosity substrate is about 1 mil (0.001 inch; 25.4 μm), about 2 mils (0.002 inch; 50.8 μm), about 3 mils (0.003 inch; 76.2 μm), about 4 mils (0.004 inch; 101.6 μm), about 5 mils (0.005 inch; 127 μm), about 6 mils (0.006 inch; 152.4 μm), about 7 mils (0.007 inch; 177.8 μm), and about... 8 mil (0.008 inch; 203.2 μm), about 9 mil (0.009 inch; 228.6 μm), about 10 mil (0.01 inch; 254 μm), about 12 mil (0.012 inch; 304.8 μm), about 15 mil (0.015 inch; 381 μm), about 20 mil (0.02 inch; 508 μm), about 25 mil (0.025 inch; 635 μm), or about 30 mil (0.03 inch; 762 μm).

[0088] The fibrillated submicron surface structure of the low-porosity substrate is configured to retain algae via anchorage. The depth of the fibrillated submicron surface structure and the porosity of the low-porosity substrate are sufficient to retain algae via anchorage while preventing algae from growing inward beyond the depth of the nodes and fibrillary nanostructures into the low-porosity substrate. In some embodiments, multiple algal spores are seeded onto the substrate and allowed to develop into seedlings, during which time the algae form anchorage structures on the surface of the substrate. In other embodiments, multiple seedlings (e.g., sporophytes and / or gametophytes) are directly seeded onto the substrate and allowed to form anchorages on the substrate. The multiple algal spores and / or juvenile algae may all belong to the same species, or belong to two or more different species. In some embodiments, two different algal species exhibit a symbiotic relationship when cultured or grown together.

[0089] In addition to retaining seaweed through seaweed anchorage, the culture system and substrate disclosed herein can promote the germination and growth of sown seaweed spores, as well as the growth of juvenile and mature seaweed. The culture substrate can, for example, create a microenvironment conducive to the germination and growth of sown seaweed spores, as well as the growth of juvenile and mature seaweed.

[0090] In some embodiments, the culture medium provides selective nanostructures that facilitate the formation of anchorage and subsequent growth of one or more target algal species, while inhibiting or preventing the attachment or growth of non-target species or other organisms. That is, the nanostructures of the culture medium support the attachment and growth of algal species while inhibiting biofouling. In some embodiments, when biofouling species (e.g., non-target species or other organisms) do attach to the culture medium, the adhesion is weaker than that of the target algal species, and the biofouling species can be removed, for example, by rinsing. In such embodiments, the physical removal of biofouling does not result in significant movement of the target species.

[0091] In some implementations, the culture substrate promotes rapid and healthy growth of the target species, allowing it to produce and secrete natural antifouling compounds before biofouling species can establish themselves on the substrate. Therefore, in addition to the fibrillated submicron surface structure of the low-porosity matrix, the target species also contribute to resistance to biofouling.

[0092] Selective nanostructures can be achieved, for example, by providing a combination of interfibrillary distance, matrix porosity, and depth of fibrillated submicron surface structures, which support the attachment and growth of target algal species while inhibiting or preventing the attachment and growth of biopollutants.

[0093] Good settling and attachment are crucial for the successful cultivation of seaweed crops; seedlings must attach firmly enough to prevent separation from the culture medium under the extreme exposure conditions of the open ocean. All juvenile seaweeds are susceptible to biofouling, which is usually caused by the overgrowth of other algal species, including, for example, diatoms, filamentous brown algae, and green algae. Biofouling problems are initially most prevalent on farm sites and can be small enough to pose a suffocating hazard, but it can sometimes occur in nurseries during seed production. An ideal culture medium should ensure firm attachment of the target species while inhibiting the growth of biofouling organisms.

[0094] The fibrillated submicron surface structure of the low-porosity matrix described in this article supports the strong adhesion and growth of algae while inhibiting biofouling. Figure 4 A young kelp plant 400 is depicted attached to a low-porosity matrix 450 with a fibrillated submicron surface structure. The young kelp plant 400 is attached to the low-porosity matrix 450 via a fixation 410, which appears as a network of protrusions emanating from the base of the young kelp plant 400. Figure 5 , 6As shown in the two samples on the left of each of samples 7, this attachment was observed, according to some embodiments, when kelp, nori, and red algae were seeded and grown on low-porosity substrates with fibrillated submicron surface structures, respectively. Conversely, low-porosity substrates lacking fibrillated submicron surface structures could not retain algae, such as... Figure 5 , 6 The two samples to the right of each of samples 7 are shown. The surface structure of the low-porosity matrix lacking fibrillated submicron surface structure is as follows: Figures 3A-3D As shown. Figures 3A-3D These are SEM micrographs depicting the surface structure of a low-porosity ePTFE matrix lacking surface nodes and fibrillary nanostructures. Figures 3A-3C The surface structure on the first side of the low-porosity matrix was depicted at increased magnification. The scale presented is 100 μm. Figure 3A ), 10μm ( Figure 3B ) and 5μm ( Figure 3C ). Figure 3D The surface structure on the second side of the low-porosity matrix is ​​depicted (providing a scale of 5 μm). At the lowest magnification ( Figure 3A The surface of the low-porosity matrix appears almost smooth, and is consistent with... Figure 2A The surface of the low-porosity matrix depicted in the image is similar. At higher magnifications, it becomes clear that the substrate lacks fibrillated submicron surface structures. Figure 3B-3D ).

[0095] In some embodiments, the culture medium includes a high-porosity matrix in addition to a low-porosity matrix. In some embodiments, the high-porosity matrix has a porosity of at least 30%, and a node and protofibril microstructure, characterized by an average interfibril distance of about 1 μm to about 500 μm, or an average pore size of about 1 μm to about 500 μm. The high-porosity matrix can, for example, disperse the low-porosity matrix, help control the positioning of algae on the low-porosity matrix, and transport nutrients to growing algae. In some embodiments, the high-porosity matrix can retain and support spore growth (e.g., retain and support algal spores and the growth of mature algae therefrom), and / or inhibit or prevent the retention of spores and / or biocontamination of organisms. Whether high porosity retains and supports spore growth or inhibits such retention depends on the characteristics of the high-porosity matrix, such as porosity and interfibril distance.

[0096] In some embodiments, the high-porosity matrix has a microstructure comprising a plurality of fibrils defining an average interfibril distance. Figure 8Figure 1 shows a SEM micrograph depicting the microstructure 100 of a high-porosity matrix comprising a fibrillated material according to some embodiments. The fibrillated material with microstructure 800 shown in Figure 1 is expanded polytetrafluoroethylene (ePTFE). As shown, microstructure 800 is defined by a plurality of fibrils 802 interconnected with nodes 804. The fibrils 802 define interfibril spaces 803.

[0097] The fibril 803 has a defined average interfibril distance, which in some embodiments may be about 1 μm to about 500 μm, 1 μm to about 200 μm, about 1 μm to about 50 μm, about 1 μm to about 20 μm, about 1 μm to about 10 μm, about 1 μm to about 5 μm, about 5 μm to about 50 μm, about 5 μm to about 20 μm, about 5 μm to about 10 μm, about 10 μm to about 100 μm, about 10 μm to about 75 μm, about 10 μm to... Approximately 50 μm, approximately 10 μm to approximately 25 μm, approximately 25 μm to approximately 200 μm, approximately 25 μm to approximately 150 μm, approximately 25 μm to approximately 100 μm, approximately 25 μm to approximately 50 μm, approximately 50 μm to approximately 200 μm, approximately 50 μm to approximately 150 μm, approximately 50 μm to approximately 100 μm, approximately 100 μm to approximately 200 μm, approximately 100 μm to approximately 150 μm, approximately 150 μm to approximately 200 μm, or approximately 200 μm to approximately 500 μm. In some embodiments, the average interfibrillary distance of the fibrils 802 can be about 1 μm, about 2 μm, about 3 μm, about 4 μm, about 5 μm, about 10 μm, about 20 μm, about 30 μm, about 40 μm, about 50 μm, about 60 μm, about 70 μm, about 80 μm, about 90 μm, about 100 μm, about 110 μm, about 120 μm, about 130 μm, about 140 μm, about 150 μm, about 160 μm, about 170 μm, about 180 μm, about 190 μm, about 200 μm, about 300 μm, about 400 μm, or about 500 μm.

[0098] Figure 9 yes Figure 8 The microstructure is shown in a higher magnification SEM image. Figure 9 Determine the dimensions of the selected interfiber space 803, in μm.

[0099] Figure 10 These are SEM micrographs based on some embodiments, showing another microstructure of a high-porosity matrix comprising fibrillated ePTFE material.

[0100] Figure 11 yes Figure 10 The microstructure is shown in a higher magnification SEM image.

[0101] In some embodiments, at least some protofibrils 802 are sufficiently spaced apart to retain spores in the interfibril spaces 802. In other embodiments, the protofibrils 802 are sufficiently spaced apart to inhibit or prevent spores from being retained in the interfibril spaces 802.

[0102] Figure 12 This is a perspective view of a schematic diagram of the microstructure of a culture substrate according to some embodiments. As shown, the microstructure 1200 is defined by a plurality of pores 1202.

[0103] The aperture 1202 can be circular, nearly circular, or oblong. The aperture 1202 can have a diameter of approximately 1 μm to 500 μm, approximately 1 μm to 200 μm, approximately 1 μm to 50 μm, approximately 1 μm to 20 μm, approximately 1 μm to 10 μm, approximately 1 μm to 5 μm, approximately 5 μm to 50 μm, approximately 5 μm to 20 μm, approximately 5 μm to 10 μm, approximately 10 μm to 100 μm, approximately 10 μm to 75 μm, approximately 10 μm to 50 μm, and approximately 10 μm to 25 μm. The diameter or approximate diameter is approximately 25 μm to 200 μm, approximately 25 μm to 150 μm, approximately 25 μm to 100 μm, approximately 25 μm to 50 μm, approximately 50 μm to 200 μm, approximately 50 μm to 150 μm, approximately 50 μm to 100 μm, approximately 100 μm to 200 μm, approximately 100 μm to 150 μm, approximately 150 μm to 200 μm, or approximately 200 μm to 500 μm. In some embodiments, the aperture 1202 may have a diameter or approximate diameter of about 1 μm, about 2 μm, about 3 μm, about 4 μm, about 5 μm, about 10 μm, about 20 μm, about 30 μm, about 40 μm, about 50 μm, about 60 μm, about 70 μm, about 80 μm, about 90 μm, about 100 μm, about 110 μm, about 120 μm, about 130 μm, about 140 μm, about 150 μm, about 160 μm, about 170 μm, about 180 μm, about 190 μm, about 200 μm, about 300 μm, about 400 μm, or about 500 μm.

[0104] In some implementations... Figure 8 The space between the fibrils is formed by 803 Figure 12 The pores 1202. That is, a microstructure 800 with multiple fibrils 802 can form a porous microstructure 1200. However, not all microstructures 1200 with pores 1202 are fibrillated.

[0105] In some embodiments, the microstructure of the high-porosity substrate is configured to retain spores and sporophytes, gametophytes, or other organisms growing from the retained spores. In some embodiments, the microstructure is configured to retain algal spores, algal sporophytes and / or gametophytes, plant spores, seedlings, bacterial endospores, fungal spores, or combinations thereof. In some embodiments, the culture medium retains multiple spores and / or organisms growing therefrom (e.g., sporophytes and / or gametophytes). The multiple spores and / or organisms may all be of the same type, or two or more different types. In some embodiments, the high-porosity substrate retains algal spores and / or algae of the same type seeded and attached to a low-porosity substrate. In other embodiments, the high-porosity substrate retains algal spores and / or algae of different types seeded and attached to a low-porosity substrate. In some embodiments, the culture medium retains two different spore types that exhibit a symbiotic relationship when cultured or grown together. For simplicity, the term “spore” will be used throughout this disclosure in relation to low-porosity substrates; however, gametophytes, sporophytes, seedlings or other organisms that grow from spores are also covered by the term and are considered to fall within the scope of this disclosure.

[0106] In some embodiments, in addition to preserving spores, the high-porosity matrix promotes the germination and growth of the preserved spores. That is, the high-porosity matrix can effectively maintain the preserved spores. In some embodiments, the microstructure is configured to non-removably anchor at least a portion of the spores.

[0107] For example, a highly porous matrix creates a microenvironment conducive to the germination and growth of retained spores. In some embodiments, the microstructure initially exists in a first retention phase, where it serves to retain and maintain the target spores. The microstructure then enters a second growth phase, inducing spore germination, and the spore seedlings (e.g., sporophytes, gametophytes, seedlings, etc.) that grow from the spores grow inwards onto and / or into the microstructure, resulting in mechanical coupling or anchoring of the seedlings to the microstructure. Therefore, in some embodiments, the microstructure is configured to non-removably anchor germinating spores, preventing loss during, for example, transport or placement in the field (e.g., open water environments) or due to environmental factors (e.g., water flow).

[0108] In some embodiments, the high-porosity matrix creates a selective microenvironment that favors the germination and growth of target spores while inhibiting or preventing the germination, growth, and / or proliferation of non-target spores or other cells. For example, this selective microenvironment can be achieved by providing a combination of fibril spacing and / or pore size, material density, the ratio of fibril spacing to average material density, depth or thickness, hydrophobicity, and the presence of a nutrient source, moisture, bioactive agent, and binder, which supports the germination and growth of target spores while inhibiting or preventing the germination, growth, and / or proliferation of non-target spores or other cells.

[0109] Several factors can affect the retention and / or effective maintenance of spores and the organisms that grow from them. These factors include, for example, the interfibrillary distance and / or pore size, material density, the ratio of interfibrillary distance to average material density, depth or thickness, hydrophobicity, and the presence of nutrient sources, moisture, bioactive agents, and binders. These factors will be described in more detail below.

[0110] The distance between two protofibrils (i.e., the inter-protofibril distance) defines an inter-protofibril space 803. In some embodiments, the inter-protofibril space 803 (i.e., the inter-protofibril distance) is sufficient to retain spores therein; the spores are retained between the two protofibrils defining the inter-protofibril space. The inter-protofibril distance is sufficient to allow at least a portion of the spore to enter between the two protofibrils defining the inter-protofibril space 803. In some embodiments, this thereby retains the spores within the microstructure of the culture medium. Figure 13 yes Figure 9 An improved version of the photograph shows the microstructure of a highly porous matrix containing fibrillated material and covered with exemplary spores with a diameter of about 10 μm (e.g., laver and kelp spores) or about 30 μm (e.g., red algae spores). Figure 13 This shows how and where the target spore enters between two protofibrils that define the space between the protofibrils.

[0111] In some embodiments, the average interfibrillary distance of the high-porosity matrix is ​​controlled to facilitate the entry of at least some spores into the microstructure. For example, if it is desired that the microstructure retains spores of *Palmariapalmate* with a diameter of about 30 μm, the average interfibrillary distance of the high-porosity matrix microstructure is about 30 μm or slightly larger (e.g., about 32 μm to about 35 μm). If it is desired that the high-porosity microstructure retains spores of *Porphyra* and *Kelp* with spore diameters of about 10 μm each, the average interfibrillary distance of the microstructure is about 10 μm or slightly larger (e.g., about 12 μm to about 15 μm). In some embodiments, it may be desirable to retain spores of multiple species (e.g., *Porphyra*, *Porphyra*, and *Kelp*). In these embodiments, the average interfibrillary distance is sufficient to allow at least a portion of the spores of multiple species to enter and remain in the interfibrillary space. In some embodiments, the target spore has a diameter of about 0.5 μm to about 200 μm.

[0112] In some embodiments, approximately half of the target spore can enter the interfibrillary space 803 within the highly porous matrix. In these embodiments, the interfibrillary distance is at least equal to the size of the target spore (e.g., diameter or width). In some embodiments, the interfibrillary distance is slightly larger than the size of the target spore. This allows the entire spore to enter and remain within the interfibrillary space 803.

[0113] In some embodiments, more than half of the target spores can enter the interfibrillary space 803 of the high-porosity matrix, and at most the entire spore can enter. In these embodiments, the portion of the spore entering the interfibrillary space 803 can be controlled by the depth of the pore, the opening of which is defined by the interfibrillary space. The depth of the pore can be controlled, for example, by the material density.

[0114] In some embodiments, only a portion of the spores enters the interfibrillary space 803 of the highly porous matrix. Therefore, when the interfibrillary distance is less than the diameter of the target spore, the target spore can only partially enter the interfibrillary space 803. Even when only a portion of the target spores enters the interfibrillary space 803, the target spores can still be retained therein if sufficient portion of the target spores enters the interfibrillary space 803. In some embodiments, substances applied to the microstructure, such as adhesives, can reduce the amount of spores required to enter the interfibrillary space 803 and help with retention.

[0115] In some embodiments, the microstructure of the high-porosity matrix is ​​formed from a non-fibrillated material. In some embodiments, the pore openings 1202 are inherent to the material used to cultivate the matrix. It is understood that different materials can have different pore opening characteristics, and materials can be manufactured or otherwise processed to provide the desired pore opening characteristics. In other embodiments, the pore openings 1202 are formed by micro-drilling techniques, such as: mechanical micro-drilling, such as ultrasonic drilling, powder spraying, or abrasive waterjet machining (AWJM); thermal micro-drilling, such as laser machining; chemical micro-drilling, including wet etching, deep reactive ion etching (DRIE), or plasma etching; and hybrid micro-drilling techniques, such as spark-assisted chemical engraving (SACE), vibration-assisted micromachining, laser-induced plasma micromachining (LIPMM), and water-assisted micromachining.

[0116] In embodiments where the microstructure of the high-porosity matrix is ​​formed of a non-fibrillated material, the pore opening 1202 functions very similarly to the interfibrillary space 103 and has a sufficient size to allow at least a portion of the target spores to enter the pore opening 1202. In some embodiments, this retains the spores within the microstructure of the culture substrate. In some embodiments, the size of the pore opening 1202 is controlled to facilitate at least a portion of the target spores entering the microstructure. For example, if it is desired that the microstructure of the high-porosity matrix retains spores of *Palmariapalmate* with a diameter of approximately 30 μm, the diameter of the pore opening 1202 of the microstructure is approximately 30 μm or slightly larger (e.g., approximately 32 μm to approximately 35 μm). In some embodiments, the target spores have a diameter of approximately 0.5 μm to approximately 200 μm.

[0117] In some embodiments, approximately half of the target spore can enter the pore opening 1202 in the high-porosity matrix. In these embodiments, the pore opening is at least equal to the size of the target spore (e.g., diameter or width). In some embodiments, the pore opening is slightly larger than the size of the target spore. This allows the entire spore to enter and remain within the pore opening 1202.

[0118] In some embodiments, more than half of the target spores can enter the pore opening 1202 of the high-porosity matrix, and at most the entire spore can enter. In these embodiments, the portion of the spore entering the pore opening 1202 can be controlled by the pore depth. The pore depth can be controlled, for example, by the material density.

[0119] In some embodiments, only a portion of the spores enters the pore opening 1202. Therefore, when the pore opening is smaller than the diameter of the target spore, the target spore can only partially enter the pore opening 1202. Even when only a portion of the target spores enters the pore opening 1202, the target spores can still be retained therein when a sufficient portion of the target spores enters the pore opening. In some embodiments, substances applied to the microstructure, such as adhesives, can reduce the amount of spore required to enter the pore opening 1202 and help with retention.

[0120] In some embodiments, the high-porosity matrix is ​​a low-density material. The low-density material can be fibrillated or non-fibrillated, and in some embodiments, the microstructure of the culture substrate is defined. The density of the low-density material can be about 0.1 g / cm³. 3 Approximately 0.2 g / cm³ 3 Approximately 0.3 g / cm³ 3 Approximately 0.4 g / cm³ 3 Approximately 0.5 g / cm 3 Approximately 0.6 g / cm³ 3 Approximately 0.7 g / cm³ 3 Approximately 0.8 g / cm³ 3 Approximately 0.9 g / cm³ 3 Or approximately 1.0 g / cm³ 3 In some embodiments, the density of the low-density material is approximately 0.1 g / cm³. 3 Approximately 1 g / cm 3 .

[0121] In some embodiments, the low-density material provides sufficient pore depth to retain spores in the interfibrillary space 803 or pore opening 1202.

[0122] In some implementations, the size of the pore opening (length (μm) and width (μm)) (whether formed of fibrillated or non-fibrillated material) along with the depth (μm) of the target spore entering the pore defines the capture rate. The capture rate required to adequately retain cells through the microstructure of a high-porosity matrix varies for each spore type. The required capture rate may be influenced by the properties of the materials constituting the microstructure of the high-porosity matrix and the presence of nutrients, binders, and / or bioactive agents.

[0123] In some implementations, the low-density material allows spores to germinate and grow into the low-density material. For example, when *Rhodophyta erythrodermic* spores retained in a low-density material having the microstructure described herein develop into gametophytes and then into sporophytes, *Rhodophyta erythrodermic* grows into the low-density material in all three dimensions (i.e., in the horizontal x and y dimensions and the depth z-dimensional dimension). This three-dimensional growth improves the retention of *Rhodophyta erythrodermic* gametophytes and sporophytes.

[0124] Figure 14Aand 14B These are cross-sectional SEM micrographs of low-density, high-porosity microstructured materials taken at two different magnifications according to some embodiments, showing the three-dimensional inward growth of red algae into the low-density material. Figure 14C These are cross-sectional micrographs produced using optical fluorescence microscopy, showing the red algae growing inward into low-density material.

[0125] Figure 15 (Above) is a SEM micrograph of the surface of a low-density, high-porosity microstructured material according to some embodiments. Figure 15 (The image below) shows the same culture substrate as the image above, but after the spores of kelp have been sown and germinated.

[0126] Figure 16 The images show SEM micrographs of the microstructured surface taken at two different magnifications, in which the red algae can be clearly seen attaching to and growing into the microstructure. Figure 17 The image shows a fluorescence micrograph of the surface of the microstructure, onto which red algae attach and grow. The algae are observed to grow into the microstructure in all three dimensions in a "growth network" manner.

[0127] from Figure 14A-17 The micrographs clearly show that the red algae can grow into the microstructure of the fibrillated high-porosity ePTFE matrix in all three dimensions, firmly fixing the algae in the microstructure.

[0128] In some embodiments, germinating spores grow deep into the microstructure of a highly porous matrix. This deep inward growth and integration into the microstructure provides the additional benefit of protecting the germinating spores from external environmental influences (e.g., the influence of the ocean and its currents in the case of algal gametophytes). In some embodiments, the penetration depth of the germinating spores is approximately 1:1 to approximately 200:1 relative to the initial size of the spore. For example, for a *Rhodophyta* spore with an initial diameter of approximately 30 μm, the *Rhodophyta* sporophyte can grow into the microstructure to a depth of approximately 30 μm to approximately 6 mm.

[0129] In some embodiments, the low-density, high-porosity material has a thickness sufficient to allow for the desired level of inward growth. In some embodiments, the culture substrate comprises a single layer of low-density material. In some embodiments, the culture substrate comprises two or more layers of low-density material. In some embodiments, two or more layers are present in a laminate, i.e., a laminate of multiple layers of low-density material.

[0130] In some embodiments, the interfibrillary distance and density of the microstructured high-porosity material define the average interfibrillary distance (μm) and average density (g / cm³) of the fibrillated material.3 The ratio of the average interfibrillary distance (μm) to the average density (g / cm³) of the fibrillated material. In some embodiments, the ratio is... 3 The ratio can be approximately 1:1, 10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 100:1, 125:1, 150:1, 175:1, 200:1, 225:1, 250:1, 275:1, 300:1, 325:1, or approximately 3 50:1, approximately 375:1, approximately 400:1, approximately 425:1, approximately 450:1, approximately 475:1, approximately 500:1, approximately 550:1, approximately 600:1, approximately 650:1, approximately 700:1, approximately 750:1, approximately 800:1, approximately 900:1, approximately 1000:1, approximately 1250:1, approximately 1500:1, approximately 1750:1, or approximately 2000:1. In some embodiments, the average interfibrillary distance (μm) and average density (g / cm³) of the fibrillated material are... 3 The ratio is approximately 1:1 to approximately 2000:1.

[0131] In some embodiments, the culture medium (i.e., a low-porosity matrix, a high-porosity matrix, or both) includes one or more adhesives. The adhesive may be applied to the surface of a fibrillated submicron surface structure, the surface of a low-porosity matrix, or the surface of a microstructure of a high-porosity matrix, and absorbed within the low-porosity or high-porosity matrix; or it may be applied to the surface of a fibrillated submicron surface structure, the surface of a low-porosity matrix, or the surface of a microstructure of a high-porosity matrix and absorbed within the low-porosity or high-porosity matrix. In some embodiments, the adhesive includes one or more cell adhesive ligands specific to spores to be retained by the culture medium.

[0132] In some embodiments, the culture medium described herein (i.e., a low-porosity substrate, a high-porosity substrate, or both) includes a nutrient phase incorporated with at least a portion of the culture medium. The nutrient phase serves to effectively maintain spores, germinating spores retained in the culture medium, and growing organisms (e.g., juvenile algae). In some embodiments, the nutrient phase promotes the germination and growth of spores retained within the microstructure of the high-porosity substrate. In some embodiments, the nutrient phase serves to maintain and / or promote attachment to the low-porosity substrate and / or the high-porosity substrate, or to maintain and / or promote inward growth or integration into the microstructure of the high-porosity substrate.

[0133] In some embodiments, the nutrient phase acts as a chemical attractant, capable of attracting spores or juvenile organisms (e.g., algal sporophytes and / or gametophytes) to a predetermined location on a culture medium in which the nutrient phase is applied or includes the nutrient phase.

[0134] The nutrient phase can be contained as a filler in a low-porosity matrix, on a fibrillated submicron surface structure of a low-porosity matrix, within a microstructure of a high-porosity matrix, on a microstructure of a high-porosity matrix (e.g., on its surface), or any combination thereof. In some embodiments, the nutrient phase is applied as a coating to the surface of the culture substrate. In some embodiments, the nutrient phase is contained in one or more materials forming the culture substrate. When the nutrient phase is contained in a material forming a low-porosity matrix, it can promote attachment and fixation development. When the nutrient phase is contained in a material forming a high-porosity matrix, it can promote inward growth or integration into the microstructure. By promoting the growth of algae, the nutrient phase can help prevent biofouling, as healthy, rapidly growing algae are known to produce and release their own natural antifouling compounds.

[0135] In some embodiments, the nutrient phase includes at least one nutrient that is beneficial to the attachment of the target algal species and / or target spores and the resulting germinating spores to or retention in the culture medium. For example, when *Rhodophyta* attaches to the fibrillated submicron surface structure of a low-porosity matrix or is retained by the microstructure of a high-porosity matrix, the nutrient phase may include macronutrients (e.g., nitrogen, phosphorus, carbon, etc.), micronutrients (e.g., iron, zinc, copper, manganese, molybdenum, etc.), and vitamins (e.g., vitamin B12, thiamine, biotin) that support the growth and health of germinating *Rhodophyta* spores. The nutrients in the nutrient phase can be provided in various forms. For example, nitrogen can be provided in the form of ammonium nitrate (NH4NO3), ammonium sulfate ((NH4)2SO4), calcium nitrate (Ca(NO3)2), potassium nitrate (KNO3), urea (CO(NH2)2), etc. Those skilled in the art will recognize which nutrients are beneficial to include in the nutrient phase to effectively maintain the retention of spores and the resulting germinating spores in the culture medium.

[0136] The nutrients included in the nutrient phase will depend on which spores the culture medium retains, as different spore types, germinating spores, and growing organisms (e.g., algae) will have different nutritional requirements. The choice of nutrients may also depend on the intended use of the culture system. For example, if a culture medium retaining spores, germinating spores, and / or growing organisms is introduced into an environment lacking essential nutrients, then all necessary nutrients can be included in the nutrient phase. If a culture medium retaining spores / germinating spores / growing organisms is introduced into an environment with at least one essential nutrient, then those environmentally available essential nutrients can be excluded from the nutrient phase or included in the nutrient phase at a lower concentration. The culture medium can also concentrate nutrients from the environment by trapping environmental nutrients in the microstructure, for example, a highly porous matrix. This can be advantageous in environments where environmental nutrients are present only at low concentrations.

[0137] In some embodiments, as further described elsewhere herein, the culture system can be used to transport preserved spores / germinating spores from one location to another. When the culture system functions as a transport system, the nutrient phase may include sufficient nutrient levels to effectively support the preserved spores / germinating spores / growing organisms during transport. In some embodiments, the nutrient phase may include sufficient nutrient levels to effectively maintain the preserved spores / germinating spores / growing organisms after transport, following their introduction into a new environment (e.g., open water).

[0138] In some embodiments, the nutrient phase includes one or more carriers. The carrier may include, for example, liquid carriers, gel carriers, and hydrogel carriers. In some embodiments, the carrier of the nutrient phase is an adhesive. Including an adhesive as a carrier for the nutrient phase ensures that the nutrient phase remains on and / or within the culture medium. When the nutrient phase is applied to the surface of the culture medium and includes an adhesive as a carrier, the nutrient surface can also promote adhesion to the culture medium.

[0139] In some implementations, the nutrient phase is formulated to control the release rate of nutrients.

[0140] In some embodiments, the culture medium further comprises a salt bound to it. In some embodiments, the salt is sodium chloride (NaCl). The salt bound to the culture medium can create and maintain a saline microenvironment for the retained spores / germinating spores. This can be particularly advantageous when algae and marine plants are retained in the culture medium. In some embodiments, the saline microenvironment within the culture medium can be maintained when it is immersed in fresh water, thereby effectively maintaining marine species and avoiding the need to maintain a saline culture environment, which is difficult and expensive.

[0141] In some embodiments, the culture substrate includes a liquid phase incorporated with at least a portion of the culture substrate. The liquid phase is used to provide and maintain moisture in the microenvironment of the microstructure of the high-porosity matrix, which may be beneficial for effectively maintaining spores / germinating spores / growing organisms retained by the culture substrate.

[0142] In some embodiments, the culture medium includes a liquid wicking material. The liquid wicking material can be the same material used to form low-porosity and / or high-porosity matrices. The function of the liquid wicking material is to retain moisture within the microenvironment of the culture medium.

[0143] While spores and endospores can be efficiently maintained in arid environments, germinating spores and growing organisms (e.g., juvenile algae) typically require water to grow and / or proliferate. By maintaining a moist microenvironment (e.g., by including a liquid-containing substrate and / or liquid wicking material), culture systems in which spores / germinating spores / growing organisms are retained can be transported without having to keep the culture system in an aqueous environment.

[0144] In some embodiments, the liquid phase is entrained in a low-porosity matrix, on a fibrillated submicron surface structure of a low-porosity matrix, within a microstructure of a high-porosity matrix, on a microstructure of a high-porosity matrix (e.g., on its surface), or any combination thereof. In some embodiments, the liquid phase is applied as a coating to the surface of the culture medium. In some embodiments, the liquid phase is contained in one or more materials forming the culture medium.

[0145] In some embodiments, the liquid phase includes, for example, hydrogels, slurries, pastes, or combinations of hydrogels, slurries, and / or pastes. In some embodiments, the liquid phase is a carrier of the nutrient phase.

[0146] In some embodiments, at least a portion of the culture medium is hydrophilic. This hydrophilic portion of the culture medium may facilitate retention by the culture medium and / or adhesion to the culture medium.

[0147] In some embodiments, at least a portion of the culture medium is hydrophobic. This hydrophobic portion of the culture medium can reduce, prevent, or resist the retention and / or attachment of spores / germinating spores / growing organisms. This may help reduce or prevent biocontamination and unwanted spores or other cells or organisms from attaching to the culture medium.

[0148] In some embodiments, one or more portions of the culture medium are hydrophobic and one or more portions of the culture medium are hydrophilic, thereby selectively promoting the retention of spores / germinating spores / growing organisms by or attachment to one or more hydrophilic portions of the culture medium.

[0149] In some embodiments, the culture medium may include one or more bioactive agents bound to it. Bioactive agents include any agent that has a positive or negative effect on cells or organisms in contact with the agent. Suitable bioactive agents may include, for example, biocides and serum. Biocides may be bound to portions of the culture medium to prevent unwanted cells or organisms from attaching to and growing on those portions. For example, unwanted cells may include non-target cells such as bacteria, yeast, and algae (i.e., biological contaminants). Biocides may also deter pests, such as insects. In some embodiments, biocides prevent target spores from attaching to and growing on unwanted portions of the culture medium. In some embodiments, serum may be applied to portions of the culture medium. Serum promotes spore attachment and retention and / or promotes spore germination or growth. Serum may include, for example, cell adhesion ligands, and provide a source of growth factors, hormones, and attachment factors.

[0150] In some embodiments, the culture medium is patterned. By patterning the culture medium, the regions of the culture medium on which target spores / germinating spores / growing organisms (e.g., juvenile algae) attach can be specified. In some embodiments, the culture medium includes patterns of low-porosity matrix portions and high-porosity matrix portions. In some embodiments, the culture medium is patterned in a "checkerboard" manner, having alternating portions of low-porosity and high-porosity matrix. The patterns of the low-porosity and high-porosity matrix can be organized or selective, or they can be random. The portions of the matrix can all have the same size or different sizes. A portion of the low-porosity or high-porosity matrix can have the same size but different sizes from other portions (i.e., all portions of the low-porosity matrix have the same size, but different sizes from the portions of the high-porosity matrix).

[0151] In some embodiments, the fibrillated submicron surface structure of the low-porosity matrix and / or the microstructure of the high-porosity matrix are patterned. By specifically patterning the fibrillated submicron surface structure, the microstructure, or both, target spores can be specifically retained in said portions of the microstructure while excluding cells from other portions.

[0152] In some embodiments, the fibrillated submicron surface structure includes patterns of different surface structures. For example, the average interfibrillary distance can vary throughout the culture medium. In some embodiments, the low-porosity matrix includes patterns of portions with larger and smaller interfibrillary distances. In these embodiments, the difference in interfibrillary distances can promote the attachment and fixation development of different algal species. In other embodiments, the fibrillated submicron surface structure can be removed from certain areas, leaving a smooth surface where algae will not attach. In such embodiments, this allows for control over where algae will attach to the culture medium, particularly the low-porosity matrix.

[0153] In some embodiments, the depth of the fibrillated submicron surface structure can vary throughout the culture substrate. In some embodiments, the low-porosity matrix comprises a pattern of larger and smaller fibrillated submicron surface structure depths. In these embodiments, the depth variation of the fibrillated submicron surface structure can promote the attachment and fixation development of different algal species.

[0154] In some implementations, the interfibrillation distance and depth of the fibrillated submicron surface structure can be varied. In such implementations, the fibrillated submicron surface structure can be fine-tuned for a given application.

[0155] In some embodiments, the microstructure of the high-porosity matrix contained in the culture medium comprises a pattern of higher-density and lower-density portions. In such a configuration, the lower-density portions correspond to a portion of the microstructure configured to retain and effectively maintain the target spores, while the higher-density portions inhibit or prevent cell retention. The density pattern can extend in any dimension. For example, a high-density / low-density pattern can extend in the x or y dimension of the culture medium, or in the z dimension. When extending in the z dimension, the outermost portion is typically configured as a lower-density portion to retain and effectively maintain the target spores. The lower portions can have a higher density, or they can have a lower density than the outermost portion. If the lower portions have a higher density, it inhibits or prevents the inward growth of germinating spores. If the lower portions have a lower density than the outermost portion, it encourages and / or promotes the inward growth of germinating spores. In some embodiments, the density pattern or gradient in the z dimension is generated by concentric encapsulation of microstructure materials with different densities or by a laminated structure in which each thin layer has a different density. In some embodiments, the density pattern can extend in two or all three dimensions. In some embodiments, portions of the microstructure have density gradients.

[0156] Density can be measured in various ways, including, for example, measuring the dimensions and weight of a material. Additionally, a wetting test can be performed to obtain a density value. For example, density can be altered by changing the interfibrillary distance, the number of fibrils per unit volume, the number of pores per unit volume, and the pore size.

[0157] In some embodiments, the density of the high-porosity matrix is ​​the density of the material itself that forms the high-porosity matrix; that is, it does not contain any inclusions, such as nutrient phases, liquid phases, etc.

[0158] In some embodiments, the density of the high-porosity matrix is ​​the density of the high-porosity matrix material and its inclusions, such as a nutrient phase, a liquid phase, or a density-altering filler. In some embodiments, portions of the microstructure are filled with filler to alter the density, thereby changing the ability of those portions of the microstructure to retain spores and / or prevent inward growth into the microstructure of the high-porosity matrix.

[0159] In some embodiments, the high-porosity matrix has a pattern of higher porosity portions and lower porosity portions. In some embodiments, the lower porosity portions correspond to portions of the high-porosity matrix configured to retain and effectively maintain target spores. In some embodiments, the higher porosity portions correspond to microstructural portions configured to retain and effectively maintain target spores.

[0160] In some embodiments, the high-porosity matrix includes a pattern of larger and smaller interfibrillary distances. In some embodiments, the smaller interfibrillary distances correspond to microstructural portions configured to retain and effectively maintain spores. In these embodiments, the larger interfibrillary distances are too large to retain target spores. In other embodiments, the larger interfibrillary distances correspond to microstructural portions configured to retain and effectively maintain spores. In these embodiments, the smaller interfibrillary distances are too small to retain target spores.

[0161] In some embodiments, the pattern of the high-porosity matrix is ​​generated by controlling at least two of density, porosity, and average interfibrillary distance. In some embodiments, the pattern of the high-porosity matrix, whether involving density, porosity, average interfibrillary distance, or a combination thereof, can be an organized or selective pattern, or it can be a random pattern.

[0162] In some embodiments, the pattern of the high-porosity matrix can be set or adjusted by selectively applying longitudinal tension. Setting or adjusting the pattern by applying longitudinal tension allows for mechanical modification of the pattern. In some embodiments, the pattern in fibrillated high-porosity materials can be set or adjusted by selectively applying longitudinal tension.

[0163] In some embodiments, the patterned high-porosity matrix includes portions having two or more features that are conducive to spore retention. For example, the patterned high-porosity matrix may have a low density (i.e., about 1.0 g / cm³). 3 The same portion (or smaller) and the average interfibrillary distance selected for retaining the target spores (e.g., about 30 μm for red algae spores). These same portions can also be hydrophilic and / or include one or more of a nutrient phase, binder, and bioactive agent. For example, density, interfibrillary distance, hydrophobicity, nutrient phase, binder, and bioactive agent can be selected individually to preferentially retain the target spores.

[0164] In some embodiments, the culture medium is configured as fibers, membranes, woven products, nonwoven products, braided products, fabrics, knitted products, particulate dispersions, or combinations thereof.

[0165] In some embodiments, the culture system includes at least one of a backing layer, a carrier layer, a laminate of multiple layers, a composite material, or a combination thereof. The culture medium (i.e., a low-porosity matrix and / or a high-porosity matrix) may be deposited on the backing layer or the carrier layer, or contained within the laminate. For example, the backing layer may be a rope or wire. For instance, the culture medium may be deposited on a rope or wire to create seed ropes, thus eliminating the need for on-site seed-lined rope cultivation of seaweed in open water.

[0166] In some embodiments, the culture medium has sufficient strength to act as a conveyor belt moving through the various growth stages of the retained spores (including the harvesting of germinating spores). In some embodiments, the culture medium is deposited on a backing layer, a carrier layer, or forms a laminate to produce a culture system with sufficient strength to allow it to act as a conveyor belt moving through the various growth stages of the retained spores (including the harvesting of germinating spores).

[0167] In some embodiments, the culture medium is configured as a particulate dispersion. The fibrillated submicron surface structure of a low-porosity matrix and the microstructure of a high-porosity matrix, when pre-defined, are provided by a plurality of particles in the dispersion, which is formulated for deposition onto a backing layer or carrier matrix to form a culture system. For example, the particles can be shredded or otherwise fragmented pieces of fibers, membranes, woven, nonwoven, braided, fabric, or knitted materials having the fibrillated submicron surface structure or microstructure described herein. In some embodiments, spores contact the particles before deposition onto the backing layer or carrier matrix. In other embodiments, spores contact the particles after deposition onto the backing layer or carrier matrix. The particulate dispersion can be deposited onto the backing layer or carrier matrix by, for example, spraying, dipping, brushing, or other coating methods. In embodiments where spores contact the particles before deposition onto the backing layer or carrier matrix, care must be taken to ensure that the deposition method does not negatively affect the retained spores. In this way, spores and endospores can be more resilient and withstand deposition.

[0168] In some embodiments, the culture medium comprises an expanded fluoropolymer. In some embodiments, the expanded fluoropolymer is selected from the group consisting of expanded fluorinated ethylene propylene (eFEP), porous perfluoroalkoxyalkane (PFA), expanded ethylene tetrafluoroethylene (eETFE), expanded vinylidene fluoride-co-tetrafluoroethylene or trifluoroethylene polymer (eVDF-co-(TFE or TrFE)), expanded polytetrafluoroethylene (ePTFE), and modified ePTFE. Examples of suitable expanded fluoropolymers include fluorinated ethylene propylene (FEP), porous perfluoroalkoxyalkane (PFA), polyester sulfone (PES), poly(p-xylene) (ePPX) taught in U.S. Patent Publication No. 2016 / 0032069, ultra-high molecular weight polyethylene (eUHMWPE) taught in U.S. Patent No. 9,926,416 of Sbriglia, ethylene tetrafluoroethylene (eETFE) described in U.S. Patent No. 9,932,429 of Sbriglia, polylactic acid (ePLLA) described in U.S. Patent No. 7,932,184 of Sbriglia et al., and vinylidene fluoride-co-tetrafluoroethylene or trifluoroethylene [VDF-co-(TFE or TrFE)] polymers taught in U.S. Patent No. 9,441,088 of Sbriglia.

[0169] In some embodiments, the expanded fluoropolymer includes a nutrient phase. This can be achieved by blending the nutrient phase with the fluoropolymer resin prior to extrusion and expansion of the fluoropolymer.

[0170] In some embodiments, the culture medium comprises an expanded thermoplastic polymer. In some embodiments, the expanded thermoplastic polymer forms the microstructure of the culture medium. In some embodiments, the expanded thermoplastic polymer is selected from the group consisting of expanded polyester sulfone (ePES), expanded ultra-high molecular weight polyethylene (eUHMWPE), expanded polylactic acid (ePLA), and expanded polyethylene (ePE).

[0171] In some embodiments, the culture medium comprises an expanded polymer. In some embodiments, the expanded polymer forms the microstructure of the culture medium. In some embodiments, the expanded polymer is expanded polyurethane (ePU).

[0172] In some embodiments, the expanded polymer includes a nutrient phase. This can be achieved by blending the nutrient phase with a fluoropolymer resin prior to polymer expansion.

[0173] In some embodiments, the culture medium comprises a polymer formed by expanded chemical vapor deposition (CVD). In some embodiments, the polymer formed by expanded CVD forms the microstructure of the culture medium. In some embodiments, the polymer formed by expanded CVD is poly(p-xylene) (ePPX).

[0174] It will be recognized that some of these materials are better suited for use as either low-porosity or high-porosity matrices. In other embodiments, the low-porosity and high-porosity matrices are formed from the same type of material, but the materials can be treated differently to provide the various fibrillated submicron surface structures and microstructures described herein.

[0175] In some embodiments, the material is formed or can be treated to form a fibrillated submicron surface structure. In some embodiments, the fibrillated submicron surface structure is produced by densifying and stretching an expanded fluoropolymer, expanded thermoplastic polymer, expanded polymer, or ePPX. U.S. Patent No. 7,521,010 by Kennedy teaches examples of suitable densified fluoropolymer materials for use as low-porosity matrices, the contents of which are incorporated herein by reference in their entirety. Kennedy teaches a densified fluoropolymer article having a water vapor permeability of about 0.015 g-mm / m. 2 / day or less, and a matrix tensile strength of at least 10,000 psi in both orthogonal directions. Articles are prepared by compressing expanded porous PTFE under pressure, temperature, and time that results in near-complete elimination of porosity, followed by stretching at a temperature above the crystallization melt temperature. The stretching step results in a densified ePTFE sheet with a higher tensile strength in the stretching direction than the compressed precursor used to manufacture it.

[0176] As Kennedy taught, the ePTFE sheet or film is produced according to the teachings of U.S. Patent No. 3,953,566. The ePTFE film is then compressed according to the teachings of U.S. Patent No. 5,374,473. The densified film is then stretched at a temperature exceeding the crystallization melting temperature of PTFE. At a stretching rate of, for example, 5% per second, the stretching ratio is 12:1. The stretching process can be performed in either direction, or sequentially or simultaneously in both directions, using a scaling machine or continuously on a tenter frame or similar machine. The thickness of the compression precursor directly affects the ability to achieve high stretching, as the bulk density increases when the compression precursor is stretched at a temperature above the crystallization melting temperature of ePTFE. Stretching results in a reduction in both weight and thickness per unit area. A significant increase in the matrix tensile strength of one or more sheets is also observed. The result of densification and stretching is an extremely thin, high-PTFE bulk density film with low porosity, an unexpectedly high water vapor permeability coefficient, and high tensile strength in the x and y directions. In some embodiments, the ePTFE film is sintered prior to the densification step. Furthermore, biaxial ePTFE films may comprise two or more layers of ePTFE. The process can be carried out in a continuous manner.

[0177] In some implementations, a similar process can be applied to other expanded fluoropolymers, expanded thermoplastic polymers, expanded polymers, or ePPX to generate a dense film with a fibrillated submicron surface structure, which can be used as a low-porosity matrix in the culture medium.

[0178] In some embodiments, the densified film is stretched at a temperature below the melting temperature of the expanded fluoropolymer film. In some embodiments, the densified film is stretched at a temperature just below the melting temperature. The interfibrillary spacing and general morphology of the fibrillated submicron surface structure can be controlled by the stretching rate and stretching temperature. In some embodiments, the stretching rate and temperature are selected to produce nodes and fibrillary nanostructures, wherein the fibrils are interconnected by nodes. In other embodiments, the stretching rate and temperature are selected to eliminate node formation or minimize node formation as much as possible.

[0179] In some embodiments, the fibrillated submicron surface structure appears only on one side of the low-porosity matrix, thus providing seaweed attachment only on the side with the submicron surface structure. In some embodiments, the side without the submicron surface structure is bonded to, for example, a backing layer.

[0180] The depth of fibrillated submicron surface structures can be controlled through densification and stretching steps.

[0181] In some implementations, the fluoropolymer expands to form the microstructure of the culture medium.

[0182] In some embodiments, the culture system described herein can be used for seaweed cultivation. Seaweed spores are contacted with a culture medium under predetermined conditions for a sufficient duration, the culture medium having properties necessary to retain and effectively maintain the spores, until at least some spores germinate and are retained (i.e., attached) by the culture medium. In some embodiments, the culture medium can be incubated in a medium conducive to spore germination and the growth of germinating spores. In other embodiments, the culture system itself provides a microenvironment favorable to spore germination and the growth of germinating spores, at least for a period of time (e.g., during temporary transport).

[0183] In some embodiments, the culture medium described herein can be used as a growth substrate for the growth of spores into multicellular organisms. For example, the culture medium can be used to support the growth of algae from spores to mature algae. In some embodiments, spores to be matured into multicellular organisms are contacted with the culture medium for a sufficient time under predetermined conditions until at least some spores germinate and are retained by the culture medium.

[0184] In some implementations, seaweed spores are introduced onto a culture medium, allowing gametophytes and sporophytes to mature in a manner similar to conventional culture lines. This bypasses the traditional step of winding the culture line onto a rope, cable, or other on-site support by depositing the culture medium, with or without spores, onto it. This can be achieved when the culture medium is provided by multiple particles in a dispersion.

[0185] In other embodiments, algal sporophytes and / or gametophytes are introduced directly onto the culture substrate. Compared to spore sowing, this direct sowing can reduce the laboratory time required to generate culture lines.

[0186] Traditionally, culture lines are maintained and cultivated in a laboratory environment using sterile seawater. By incorporating sufficient salt into the microstructure of a high-porosity matrix, and by providing a saline microenvironment within the microstructure, the culture system of this application avoids the expensive and cumbersome systems required for sterile seawater circulation. In some embodiments, the seeded culture medium is maintained in a standard seaweed culture tank, where nutrients are delivered via sterile seawater. By including a nutrient phase sufficient to support seaweed growth, the need to provide external nutrients to the growing seaweed can be eliminated.

[0187] Typically, culture lines must be carefully transported in seawater while avoiding collisions to prevent gametophytes and sporophytes from detaching from the line. The culture system described in this paper allows for the safe transport of gametophytes and sporophytes without seawater. This is achieved by incorporating salt and a liquid phase into the microstructure, providing a saline microenvironment with sufficient moisture to support juvenile algae during transport.

[0188] In some implementations, by controlling the adhesion strength of seaweed to the fibrillated submicron surface structure, the culture medium or portions thereof, such as a low-porosity substrate, can be reused. By controlling the fibrillated submicron surface structure, sufficiently strong adhesion can be provided to allow cultivation, but not so strong that the adherent cannot be mechanically removed by, for example, strong washing. After removing the attached seaweed, the low-porosity substrate can be reused.

[0189] The invention of this application has been described for brevity and in conjunction with specific embodiments above. It will be apparent to those skilled in the art that various modifications and variations can be made to the embodiments described herein without departing from the spirit and scope of the invention. Therefore, the embodiments are intended to cover these modifications and variations of the invention, provided that such modifications and variations are within the scope of the appended claims and their equivalents.

Claims

1. A culture system comprising a culture substrate, the culture substrate comprising a low-porosity matrix having a porosity of 10% or less and having a fibrillated submicron surface structure configured to retain algae by immobilization, the fibrillated submicron surface structure having an average depth of 1000 nm or less, the low-porosity matrix comprising at least one of expanded polytetrafluoroethylene (ePTFE) or expanded polyethylene (ePE).

2. The culture system as described in claim 1, wherein, The fibrillated submicron surface structure is characterized by an average interfibrillary distance of up to 1000 nm and including 1000 nm.

3. The culture system as described in claim 1, wherein, The fibrillated submicron surface structure has an average depth of 5 nm or less.

4. The culture system as described in claim 1, wherein, The thickness of the low-porosity matrix ranges from 25.4 μm to 762 μm.

5. The culture system as described in claim 1, wherein, The thickness of the low-porosity matrix ranges from 25.4 μm to 127 μm.

6. The culture system as described in claim 1, wherein, The culture medium is configured as a strip, matrix (substrate), woven product, nonwoven product, braided product, knitted product, fabric, particulate dispersion, or a combination of two or more of the above.

7. The culture system as described in claim 1, wherein, The culture medium includes at least one of a backing layer, a carrier layer, a laminate of multiple layers, a composite material, or a combination thereof.

8. The culture system as described in claim 1, wherein, The PTFE matrix has a density of 0.015 g-mm / m. 2 A water vapor permeability coefficient of / day or less, and formed by a method comprising the following steps: (a) preparing a biaxially expanded PTFE film; (b) densifying the expanded PTFE film; and (c) stretching the densified expanded PTFE film.

9. The culture system of claim 8, wherein, In step (c), the densified expanded PTFE film is stretched at a temperature exceeding the crystallization melting temperature of PTFE.

10. The culture system of claim 8, wherein, The expanded PTFE film is sintered prior to step (b).

11. The culture system of claim 8, wherein, Biaxially expanded PTFE films consist of two or more layers of expanded PTFE.

12. The culture system of claim 8, wherein, Steps (a)-(c) are performed sequentially.

13. The culture system of claim 1, wherein, The culture medium further comprises a high-porosity matrix having a porosity of at least 30% and a node and fibrillary microstructure, characterized in that the average interfibrillary distance is 1 μm to 500 μm, or the average pore size is 1 μm to 500 μm.

14. The culture system of claim 13, wherein, The high-porosity matrix is ​​hydrophobic.

15. The culture system of claim 13, wherein, Low-porosity and high-porosity matrices contain the same material.

16. The culture system of claim 13, wherein, The culture medium is a patterned matrix with patterns of both low-porosity and high-porosity matrices.

17. The culture system of claim 16, wherein, The patterns of low-porosity and high-porosity matrices are organized or selective patterns.

18. The culture system of claim 16, wherein, The patterns of both low-porosity and high-porosity matrices are random patterns.

19. The culture system of claim 1, further comprising a nutrient phase combined with at least a portion of the culture system.

20. The culture system of claim 19, wherein, The nutrient phase promotes the growth of algae and / or the attachment of algae to the culture medium.

21. The culture system of claim 19, wherein, At least a portion of the nutrient phase is entrained in the culture medium, entrained on the culture medium, or entrained in both the culture medium and the culture medium.

22. The culture system of claim 19, wherein, The nutrient phase exists as a coating on the surface of the culture substrate.

23. The culture system of claim 1, wherein, The culture medium is provided by multiple particles in a dispersion that is formulated for deposition onto a backing layer or carrier matrix.

24. The culture system of claim 1, wherein, The culture medium is asymmetrical, including a protofibrotic submicron surface structure configured to retain algae on only one side.

25. A method for culturing seaweed, comprising contacting a population of seaweed gametophytes and / or sporophytes with a culture substrate of a culture system as described in any one of claims 1 to 24, until at least a portion of the seaweed gametophytes and / or sporophytes form attachments to the nanostructures of the culture substrate.

26. The method of claim 25, further comprising positioning the culture system in an open aquatic environment after a portion of the algal gametophyte and / or sporophyte population has formed an attachment to the culture substrate nanostructure.

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