DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Election/Restrictions
Applicant elected Group I encompassing claims 1-13, 16, and 17, without traverse.
Claim Status
Claims 1-13, 16-17, 19, 23-25, and 30 are pending.
Claims 1-13, and 16-17 are examined on the merits.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 10 and 12 rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 10 recites that “the mechanical stress and directional force form an isotropic singularity where all directionality is lost”. The metes and bounds of this limitation are unclear. The term “isotropic singularity” does not appear to have a commonly accepted meaning in plant biomechanics or mechanics. The ordinary meaning of “singularity “in mathematics and physics is “a point at which a mathematical function or physical quantity takes an infinite value” (Oxford Advanced Learner’s Dictionary (Oxford University Press). However, claim 10 does not indicate whether “singularity” is used in this conventional sense or in some different sense, and the specification does not expressly redefine the term or provide objective criteria by which a person of ordinary skill in the art could determine when an “isotropic singularity “has been formed.
It is unclear whether the claimed singularity requires cancellation of multiple directional forces, equal forces acting from all directions, the absence of a net force, a point at which principal stress directions cannot be distinguished, or another mechanical condition.
Although the specification discusses an “isotropic singularity” at paragraphs 0155-0157, the disclosure merely states that the normal directionality of force transmission “vanishes”, creating what “can be described” as an isotropic singularity. This language describes a proposed concept but does not expressly define the term or establish objective boundaries for the claim condition. Neither the claim nor the specification provides an objective standard or measurement by which a person of ordinary skill in the art could determine whether an “isotropic singularity” has formed or whether “all directionality” has been lost.
It is unclear whether the singularity is a mathematical point, a finite physical region, or the location of a particular cell, or a broader stress condition within the polymer material of plant tissue. The disclosure does not identify a force threshold, stress distribution, degree of directional uniformity, permissible deviation from perfect isotropy, or other test defining the claimed condition.
Moreover, paragraph 0157 acknowledges that direct tracking of principal stress distributions and trajectories in plant tissues is “virtually impossible” and that relevant mechanical parameters, including cell-wall moduli and cell turgor pressures, are largely beyond present measurement capabilities. The specification therefore does not identify a practical or objective means for determining when the claimed condition has been achieve.
Accordingly, a person of ordinary skill in the art would not be reasonably apprised of the scope of claim 10.
Claim 12 recites that “said applying mechanical stress and directional force is insensitive to environmental fluctuations”. The term “insensitive” is a relative term for which the claims and specification provide no objective standard for determining the requisite degree of insensitivity. The claim does not identify which environmental variables constitute the recited “environmental fluctuations”, the magnitude or range of such fluctuation, the property of the applied stress or force that must remain unchanged, or the amount of variation that is permitted while the force remains “insensitive”. It is unclear whether the limitation concerns fluctuations in temperature, osmotic pressure, pH, humidity, light, culture-medium composition, or another environment condition.
The specification states only the application of the mechanical stress or directional force may be “insensitive, or unresponsive, to environmental fluctuations”, without providing a quantitative or qualitative standard by which the limitation can be evaluated. Accordingly, a person of ordinary skill in the art would not be reasonably apprised of the scope of claim 12.
Claim Rejections - 35 USC § 112
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Scope of Enablement
Claims 1-13, and 16-17 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specifications, while being enabling the claimed encapsulation and culture of plant protoplasts within microspheres; does not reasonably provide enablement for the full scope of the claimed methods and products, including inducing reproductive differentiation of vegetative somatic plant cells by mechanical stress and directional force to produce reproductive germ-line plant cells, sporogenous cells, meiocytes, haploid plant cells, plant gametes, zygotes, seeds, plants, germplasm, or the reproduces. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention commensurate in scope with these claims.
An “analysis of whether a particular claim is supported by the disclosure in an application requires a determination of whether that disclosure, when filed, contained sufficient information regarding the subject matter of the claims as to enable one skilled in the pertinent art to make and use the claimed invention.” MPEP 2164.01. “A conclusion of lack of enablement means that. . . the specification, at the time the application was filed, would not have taught one skilled in the art how to make and/or use the full scope of the claimed invention [i.e. commensurate scope] without undue experimentation.” In re Wright, 999 F.2d 1557,1562, 27 USPQ2d 1510, 1513 (Fed. Cir. 1993); MPEP 2164.01.
Claim 1 is directed to an in vitro method of inducing reproductive differentiation in a vegetative somatic plant cell by applying mechanical stress and directional force to a vegetative somatic plant cell encapsulated in a polymer materials. Under the broadest reasonable interpretation, claim 1 encompasses the use of any vegetative somatic plant cell, from any plant species or tissue source, encapsulated in any polymer material, and subjected to any magnitudes, duration, spatial distribution, or type of mechanical stress and directional force sufficient to induce reproductive differentiation. Claim 2-12 depend directly or indirectly from claim 1 and therefore included all limitations of claim 1. The additional limitations relating to the polymer materials, microsphere structures, physical-property changes, external signals or forces, isotropic tension or compression, cell clusters, and environmental fluctuations do not provide the missing operative teaching for inducing reproductive differentiation in a vegetative somatic plant cell. Claim 13 is directed to a reproductive germ-line cell, haploid plant cell, or plant gamete produced by the method of claim 1 and therefore requires successful performance of the method recited in claim 1. Claims 16 and 17 further require use of the plant gamete of claim 13 to form a zygote and production of a seed, plant , or germplasm from that breeding method. Because the specification does not enable production of the cell or gamete required by claim 13, it likewise does not enable the breeding method of claim 16 or the resulting product of claim 17.
The specification provide working procedures for isolating tobacco BY-2 cell protoplasts, encapsulating the protoplasts in agarose microspheres, and further encapsulating the agarose microspheres in alginate methacrylate (pa0109-0115). The only experimentally demonstrated biological outcome is that the encapsulated protoplasts regenerated their cell walls, elongated, escaped from the agarose microspheres, and reentered normal mitotic division (pa0116). The specification does not report that any encapsulated vegetative somatic cell acquired reproductive identity, entered meiosis, formed a germ-line cell, produced a haploid cell or functional gamete, or regenerated into a plant capable of producing seed. Rather, the disclosed working example demonstrates continued vegetative growth rather than the claimed reproductive differentiation (pa0116).
Although the specification proposes that mechanically tunable polymer microenvironments may influence plant cell growth and differentiation, the disclosed double-encapsulation procedure is described as “the beginning of an in vitro culture technique” (pa0118). The specification does not establish an operative correlation between any disclosed mechanical condition and induction of reproductive differentiation. Statements that encapsulation may be used to induce reproductive differentiation, or that cells produced by the method may be haploid cells or gametes, recite the desired result but do not teach the condition required to achieve that result. Paragraph (0099) states that controlling the mechanical properties of the polymer may alter developmental fate, but the specification does not provide any experimental that demonstrating that this occurred.
The disclosed experiments likewise do not establish that the encapsulated cell undergoes reproductive differentiation solely in response to the claimed mechanical stress and directional force. Throughout the examples, the encapsulated protoplasts are maintained in media containing nutrients, plant growth regulators, ions, osmotic agents, and conditioned medium derived from BY-2 suspension cultures (pa0100-0108). Following encapsulation, the microspheres are transferred into protoplast culture medium for continued culture (pa0113-0115). The specification does not disclose that the agarose or alginate matrices prevent diffusion of soluble molecules between the surrounding culture medium and the encapsulated cell. Accordingly, the examples do not demonstrate that the alleged biological response results solely from the claimed mechanical conditions rather than continued biochemical communication with the surrounding culture environment.
To practice the full scope of claim 1, a person of ordinary skill in the art would be required to determine, without adequate guidance, at least: 1) which plant species are suitable for the claimed method; 2) which vegetative somatic cell or tissue types retain the competence to undergo reproductive differentiation; 3) whether tobacco BY-2 cells are capable of undergoing the claimed reproductive conversion; 4) whether the starting material must be a protoplast, callus cell, meristematic cell, leaf cell, or another cell type; 5) the suitable polymer composition and concentration; 6) the required polymer stiffness, elasticity, swelling properties, and degradation properties; 7) the size, shape, and amount of polymers surrounding the cell; 8) the magnitude, duration, timing, direction, and spatial distribution of the applied mechanical stress and force; 9) whether tension, compression, or a combination thereof is required; 10) whether the cell must be isolated or located within a cluster of cells; 11) the culture-medium, nutrient, and hormone conditions required to support reproductive differentiation; 12) the morphological, molecular, cytological, or functional criteria by which reproductive differentiation is identified; 13) the conditions required for meiosis, tabloidization, and formation of a functional gamete; and 14) the conditions required to obtain a zygote and viable seed from the resulting cell. Moreover, a person of ordinary skill would first have to determine whether the claimed biological phenomenon is achievable at all, because the specification does not disclose a single successful embodiment demonstrating that mechanical stress and directional force alone convert a vegetative somatic plant cell into a reproductive cell.
The specification does not identify even one operative combination of these variables that incudes reproductive differentiation. Consequently, the required experimentation would not merely involve routine optimization of a known working embodiment. Rather, a person of ordinary skill would first have to discover whether and under what conditions the proposed mechanical treatment is capable of producing the claimed biological result.
Claim 1 is not limited to a particular plant species, cell type, polymer, polymer geometry, polymer stiffness, force magnitude, force duration, culture condition, developmental stage, or type of reproductive cell. The claim therefore encompasses a large number of materially different embodiments, while the working disclosure is limited to encapsulation and somatic growth of tobacco BY-2 protoplasts.
The nature of the invention is complex and unpredictable. Ikeuchi (Momoko Ikeuchi et. al., Development (2016) 143, 1442-1451) explain that plant regeneration can involve reprogramming of differentiated somatic cells or activation of relatively undifferentiated cells, and that developmental and environmental constrains influence the regulatory mechanisms governing regeneration (Abstract and Introduction). Méndez-Hernández (Hugo A Méndez-Hernández et. al., Frontiers in Plant Science (2019) Vol10:77, pp1-15) further explain that somatic embryogenesis requires changes in the cellular gene program and involves interacting signaling pathways, plant-growth regulators, stress responses, and other factors affecting acquisition of embryogenic competence (Abstract; p3, “THE ROLE OF PLANT GROWTH REGULATORS DURING THE INDUCTION OF SOMATIC EMBRYOGENESIS”; p8 “EPIGENETICS”). These references establish that ordinary somatic growth and acquisition of a differentiated developmental fate are distinct biological outcomes governed by multiple variables. Accordingly, successful survival, cell-wall regeneration, elongation, or mitotic growth of the encapsulated tobacco BY-2 protoplasts disclosed in the specification does not , without further evidence or guidance, establish that those cells will undergo the reproductive differentiation required by the claims.
The amount of direction and guidance provided is insufficient. The specification teaches how to prepare agarose and alginate microsphere containing living plant protoplasts, but does not identify mechanical parameters or biological conditions that successfully induce reproductive differentiation. The specification provide generalized descriptions of shrink/swell polymers, isotropic tension or compression, and mechanically tunable environments, without identifying which conditions produce the claimed results.
The working examples are insufficient with respect to the claimed invention. The examples demonstrate encapsulation, survival, cell-wall regeneration, elongation, mitotic division, and double-layer microsphere formation (pa0116-0118). They do not demonstrate the claimed biological transition from a vegetative somatic cell to a reproductive cell. No experiment shows formation of an archesporial cell, sporogenous cell, meiocyte, haploid cell, egg cell, sperm cell, or functional gamete, nor do the examples demonstrate meiosis, fertilization, zygote formation, regeneration of a reproductive cell into a plant, or seed production.
The quantity of experimentation required would be extensive. A person of ordinary skill would need to screen combinations of plant species, cell sources, cell developmental stages, polymers, polymer concentrations, polymer geometries, mechanical-force conditions, culture media, hormones, and reproductive differentiation assays. Because no operative starting point is provided, such experimentation would be exploratory and amount to a research program rather than routine verification or optimization.
Although a person of ordinary skill would understand plant protoplast culture, polymer encapsulation, plant regeneration, and developmental-marker assays, the skilled person would not have been able to determine from the disclosure which mechanical and biological conditions induce the claimed reproductive differentiation without extensive and undue experimentation.
Claim 10 presents an additional enablement deficiency because it requires the mechanical stress and directional force to form an “isotropic singularity where all directionality is lost”. Even assuming this limitation can be construed, the specification does not provide a working example or sufficient technical guidance for generating, location, confirming, or reproductively using the claimed mechanical condition. The specification does not identify the required stress distribution, force magnitude, polymer geometry or material properties, duration of force application, location of the cell relative to the alleged singularity, or permissible deviation from isotropy. Specification acknowledges that tools for directly studying force transmission at the tissue level are “virtually non-existent”, that direct tracking of principal stress distributions and trajectories is “virtually impossible”, and that direct manipulation of the physical parameters governing stress distribution is extremely difficult and hard to interpret (Pa0157). This admission further demonstrates that the specification does not provide sufficient guidance for practicing the full scope of the claimed invention without undue experimentation. Thus, a person of ordinary skill would have to undertake extensive experimentation to determine how to produce and confirm the claimed condition and whether that condition induces reproductive differentiation.
Accordingly, considering the breadth of the claimed subject matter, the absence of any working embodiment that induces reproductive differentiation, the failure of the disclosed experiments to distinguish the effect of mechanical stress and directional force from continued biochemical communication with the surrounding culture environment, the unpredictability of plant-cell developmental reprogramming, the limited guidance provide regarding the necessary mechanical and biological conditions, and the extensive experimentation required to identify an operative combination of variables, the specification does not enable a person of ordinary skill in the art to make and use the full scope of the claims without undue experimentation. The disclosure teaches encapsulation and continued vegetative survival and mitotic growth of tobacco BY-2 protoplasts, but it does not teach how to convert a vegetative somatic plant cell into a reproductive germ-line cell, haploid plant cell, or functional gamete, or how to use such a cell to form a zygote, seed, plant, or germplasm as required by the claims. Claims 1-13, 16, and 17 are therefore rejected for lack of enablement.
Conclusion
No claims are allowed.
The closest teaching in the art likewise do not teach the claimed result. Grasso (Matthew S Grasso et. al., Applications in Plant Sciences 2016 4(5): 1500140, pp1-5) disclose encapsulation of living tobacco BY-2 protoplasts in agarose microbeads for handling and culture of individual plant cells. Grasso demonstrates that encapsulated protoplasts remain viable, regenerate cell walls, elongate, undergo mitotic growth, and can be maintained in culture. However, Grasso does not teach or demonstrate that encapsulation or mechanically generated force induce reproductive differentiation of vegetative somatic plant cells, conversion to reproductive germ-line cells, sprogenous cells, meiocytes, haploid cells, or functional gametes, or production of zygotes, seed, or plant. Thus, even the closest prior art established only successful encapsulation and vegetative cell growth, not the claimed mechanically induced reproductive conversion. The present specification likewise demonstrates encapsulation and vegetative growth, but does not provide the additional operative teaching required to achieve the claimed reproductive differentiation.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to YANXIN SHEN whose telephone number is (571)272-7538. The examiner can normally be reached Monday-Friday.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Amjad A Abraham can be reached at (571)272-7058. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/YANXIN SHEN/ Examiner, Art Unit 1663
/WEIHUA FAN/ Primary Examiner, Art Unit 1663
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