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 .
Priority
Applicant’s claim for the benefit of a prior-filed application no. PCT/US2022/079184 filed November 2nd, 2022, under 35 U.S.C. 119(e) or under 35 U.S.C. 120, 121, 365(c), or 386(c) is acknowledged, as is Applicant’s claim to prior-filed provisional application no. 63/274,714 filed November 2nd, 2021.
Thus, the earliest possible priority for the instant application is November 2nd, 2021.
Information Disclosure Statement
The two information disclosure statements (IDS) submitted on July 18th, 2024, were considered, initialed, and attached hereto. A signed copy of the list of references cited is included with this Office Action.
Restriction/Election
In response to the communication received on January 1, 2026, from Timothy H. Van Dyke, the election of Group II, claims 11-15, without traverse, is acknowledged. Applicants have further elected the species of PLT, without traverse. Examiner notes that during a telephone conversation on February 2nd, 2026 with Timothy H. Van Dyke, it was clarified that PLT was the intended elected species, not PLT5 as in the Remarks Made in an Amendment filed January 12th, 2026.
Status of Claims
Claims 1-20 are examined herein.
Claims 1-10 and 16-20 are withdrawn from consideration.
Claims 11-15 filed April 9th, 2025 are pending and examined herein.
Specification
The disclosure is objected to because it contains an embedded hyperlink and/or other form of browser-executable code. See pg. 24, ln. 10 for example. Applicant is required to delete the embedded hyperlink and/or other form of browser-executable code; references to websites should be limited to the top-level domain name without any prefix such as http:// or other browser-executable code. See MPEP § 608.01.
Claim Interpretation
Claim 1 recites “DR DNA”, which is undefined in the specification. This is taken to mean developmental regulator DNA, based on the use of PLT, WUS1, WIND1, and BBM/WUS proteins in the art.
Claim Objections
Claim 11 is objected to for depending on withdrawn claims (claims 1-10), as well as a cancelled claim (claim 27). As claims 11-15 depend on with drawn claim 1, it is suggested to amend claim 11 to include the vector of claim 1 to remove the dependency.
Claim 11 is objected to for lacking a preamble. It is suggested to amend similar to the following: “A method of transforming a plant to express at least one heterologous nucleic acid sequence, the method comprising…”
Claim 11 and 14 are objected to for lacking proper capitalization and italics of Agrobacteria and Agrobacterium.
In withdrawn claim 1, with examined claims 11-15 depending therefrom, “DR” is used as abbreviation, as well as “PLT”, “WUS1”, “WIND1”, and/or “BBM-WUS”. It is suggested to insert a definition for the acronyms, immediately before the first appearance of each, and to enclose the appearance of “DR”, “PLT”, “WUS1”, “WIND1”, and/or “BBM-WUS” in parentheses (at the first appearance only).
Claim 13 is objected to for lacking a period at the end of the sentence.
Appropriate correction is required.
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.
Claims 11-15 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
The claims are broadly drawn to a method of transforming any plant to express at least one heterologous nucleic acid sequence through the delivery of at least on agrobacterium comprising a vector with an expression cassette comprising at least one DR DNA sequence that encodes PLETHORA (PLT).
The instant specification reduces to practice vector construction using a PLT5 construct from Arabidopsis to improve de-novo shoot regeneration from aerial organs by wound-induced callus and Agrobacterium-mediated transformation in mature tomato and Snapdragon, wherein the Agrobacterium is suspended in an induction medium containing liquid MS consisting of 4.6 g/L MS salts and vitamins [Example 1]. The specification reduced to practice infection with Agrobacterium containing the PLT5 plasmid of cotyledon explants in sweet pepper and Bok choi tissue culture in two genotypes of Bok choi and one species of sweet pepper with successful plant regeneration and genetic transformation using MS with 3mg/L 6-BA [Example 2-3].
The specification does not reduce to practice any other PLT construct for promotion of de novo shoot regeneration. The specification does not reduce to practice all plant species in the method of transforming a plant with the vector comprising an expression cassette which comprises a sequence that encodes PLT. The specification does not reduce to practice all adult plants or all recalcitrant plant species in the method of transformation. The specification additionally does not reduce to practice any shoot, root, or callus induction medium, for the Agrobacterium suspension in induction medium.
Regarding the importance of PLT genes in plant growth, Du et al. (2017, PLETHORA transcription factors orchestrate de novo organ patterning during Arabidopsis lateral root outgrowth, PNAS, 114(44):11709-11714) teaches that PLT transcription factors are the key molecular triggers for the de novo organ patterning during Arabidopsis lateral root formation [pg. 11709, col. 2, ¶3]. Du teaches that PLT3, PLT5, and PLT7 are expressed earlier in lateral root formation, redundantly regulate the correct initiation of formative cell divisions, and are required for the activation of the PLT1, PLT2, and PLT4 genes, which contribute to stem cell maintenance and establishment of de novo meristems [pg. 11709, col. 2, ¶1]. This indicates that overexpression of any PLT may not have the same desired effect and that sufficient variety of PLT gene expression was not reflected in the instant specification.
The broad genus of “plant” encompasses the transformation of any plant, while the Applicant only reduces to practice tomato, Snapdragon, sweet pepper, and Bok choi. Tzfira et al. (2006, Agrobacterium-mediated genetic transformation of plants: biology and biotechnology, Current Opinion in Biotechnology, 17:147-154.) teaches that not all plants are readily transformable and may depend on the method of transformation, the Agrobacterium strain used, the regeneration capacity, and the plant’s own genetic makeup and defense mechanisms for efficacy of transformation [pg. 150, col. 2, ¶2].
Transformation efficiency also depends on the media used and can vary based on the nutrient balance in the medium. Pasternak (2025, Plant Tissue Culture In Vitro: A Long Journey with Lingering Challenges, Int. J. Plant Biol. 16, 97) teaches that different formulations show positive outcomes in certain plant groups; however, the applicability is species dependent [pg. 3, ¶2]. The Applicants define their root, shoot and callus induction mediums under tissue culture [pg. 25, ln. 1-4 of instant application] and only reduce to practice the use 3 mg/L 6-BA with the Bok choi and sweet pepper after they did not regenerate shoots from the calluses formed from injection with GV3101 with PLT5 [Experiments 2 and 3]. Thus, use of any shoot, root, and callus medium is not reduced to practice with the method as claimed with any plant species.
The Applicant notes that “adult plant” refers to a fully mature plant that is able to reproduce, yet concedes that, while transformation with the PLT5 construct into tomato and Snapdragon resulted in de-novo shoot regeneration and genetic transformation from wound sites on plants [Example 1 and 2], wound induction in sweet pepper and Brassica rapa ssp. chinensis with the construct promoted callus or embryo-like structure formation, but no shoot regeneration occurred [Example 3]. Examiner notes that the Applicant does not specify that the Snapdragon and tomato are adult plants, but does state that the Snapdragon plants were 70-days-old. For Bok choi, the Applicant instead tested if PLT5 promotes plant regeneration through tissue culture using cotyledons with petiole (i.e. not from adult plants), generating roots that could then be cultured on new medium. For sweet pepper, cotyledon explants or hypocotyls were infected with Agrobacterium containing PLT5 plasmid. The cotyledon explants did not regenerate shoots, but did regenerate calluses from which somatic embryos could regenerate after further culturing. The Applicant concedes that hypocotyls were unsuccessful in regenerating transgenic shoots, indicating that the effectiveness of transformation may be variable by plant tissue and by species.
The Applicant further suggests that the failure of de-novo shoot regeneration from aerial organs in Bok choi and sweet pepper was likely due to suberin in response to wounding. This suggests that any method of delivery of the Agrobacterium does not inevitably result in successful transformation of any plant, particularly transformation of an adult plant. Indeed, Luo et al. (2025, Unlocking in vitro transformation of recalcitrant plants, Trends in Plant Science, 30:12) teaches that immature embryos are commonly used for crop transformation, as juvenile explants tend to exhibit higher regenerative capacity because of the differences in phytohormone and nutrient contents [pg. 1312, ¶1]. Luo teaches that, particularly in recalcitrant plants, using fresh seeds, younger tissue, and cotyledonary nodes as explants aides in optimizing methods in recalcitrant species such as Triticum monococcum L., Bambusa arundinacea, Olea europaea, Phaseolus vulgaris, and Helianthus annuus [Table 2].
Luo teaches that within recalcitrant species, regeneration is often dependent on genotype [pg. 1313, ¶2, 4, and 6]. As the Applicant only reduced to practice tomato, snapdragon, sweet pepper, and two genotypes of Bok choi, there is not sufficient variance to represent all recalcitrant plants.
Thus, the Applicant reduces to practice a method of transformation for several recalcitrant species, but would require significant experimentation to achieve a standardized method with the explant material and necessitated hormone medium for the genus of “plants” in claim 11. The Applicant does not reduce to practice the full genus of “adult plants” of claim 12, as de-novo shoot regeneration was not possible from aerial organs in Bok choi and sweet pepper. The effectiveness of a method may vary by plant species and by induction medium, especially if the species is recalcitrant as claimed in claim 13. Additionally, PLT5 was used to represent all of the PLT family, some of which have different roles and expression times. Because of the variability within the art at the time of filing, the lack of sufficient variety of species to reflect the variance within the genus, and the lack of correlation between structure and function, the claims lack written description.
Claim Rejections - 35 USC § 102
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 11 and 13-15 are rejected under 35 U.S.C. 102(a)(2) as being anticipated by Scheres et al. (02/25/2021), “Genes for hormone-free plant regeneration,” US 20210054393 A1, (as cited in IDS filed 7/18/2024).
Claim 11 recites a method of transforming a plant to express at least one heterologous nucleic acid sequence comprising delivering to the plant at least one agrobacterium comprising the vector of any of claim 1 (withdrawn claim reciting: A vector comprising, as operably linked components, an optional first expression cassette which comprises a nucleic acid sequence encoding at least one selectable marker, an optional second expression cassette which comprises a cloning site, a third expression cassette which comprises at least one DR DNA sequence that encodes PLT, and an optional fourth expression cassette comprising a gene of interest), the vector engineered to comprise at least one transgene, wherein the delivering results in introduction of the at least one transgene into at least one plant cell of the plant such that the at least one transgene is stably integrated into a genome of said at least one plant cell, or optionally, wherein delivering to the plant at least one agrobacterium comprising the vector of any of claims 1-10 or 27 comprises delivering two or more agrobacteria, wherein an agrobacterium of the two or more agrobacteria comprises a different vector comprising a different DR DNA sequence relative to another agrobacterium of the two or more agrobacteria.
Claim 13 recites the method of claim 11, wherein the plant is a recalcitrant plant
Claim 14 recites the method of claim 11, wherein the agrobacterium is suspended in an induction media.
Claim 15 recites the method of claim 14, wherein the induction media is selected from the group consisting of callus induction media, shoot induction media, or root induction media.
Regarding claim 11 (and the vector of claim 1 from which 11 depends), Scheres discloses a method for regenerating a plant cell, preferably regenerating a shoot from a plant cell, by altering the expression levels of WOX5 and a PLT protein [Abstract]. The invention further pertains to a nucleic acid construct (used interchangeable with “vector” [¶74]) comprising at least one expression cassette and capable of containing optional expression cassettes comprising selectable markers, cloning sites, and the like [¶75], including GFP reporter genes, a CaMV35S promoter, and NPTII selectable marker [¶424], inter suitable for transient protein expression and the use of the protein combinations for regenerating a shoot from a plant cell [Abstract]. Scheres discloses that different proteins can be controlled by different types of inducible promoters which can be operably linked to one or more endogenous genes, wherein the endogenous gene encodes a protein [¶232]. Scheres discloses that the vector is introduced by Agrobacterium to deliver the transgene to the plant cell and stably integrated into the plant genome (i.e. transforming a plant to express at least one heterologous nucleic acid sequence and at least one cell of a plant has at least one transgene stably integrated into a genome of said at least one plant cell) [¶233]. For the induction of shoot regeneration from a plant cell, Scheres discloses using defined Arabidopsis transcription factors, including PLETHORA 5 (PLT5) (i.e. expression cassette comprising at least one DR DNA sequence that encodes PLT) [¶377].
Regarding claim 13, Scheres discloses that there is a need for recombinant DNA constructs that increase or induce the regenerative potential of recalcitrant plants upon introduction in the plant cell [¶8]. Scheres further discloses that, in some embodiments, the plant cell is a recalcitrant plant cell, i.e. a plant cell in which the regeneration efficiency fails or wherein the regeneration efficiency is poor, including pepper, soybean, cucumber, sugar beet, Arabidopsis, tomato, sweet pepper, and Brassica rapa [¶275 &276].
Regarding claims 14 and 15, Scheres discloses an example of tomato transformation using the transformation construct with a CaMV35S promoter and an erGFP fluorescent reporter gene electroporated to Agrobacterium tumefaciens strain GV2101 for plant transformation [¶424]. Tomato seeds were sterilized and germinated on ½MS10 medium for 11 days. Cotyledon explants from the seedlings were dissected and precultured for 24 h on MS20 medium supplemented with 40 μg.I-1 acetosyringone. The explants were submerged in a suspension of Agrobacterium tumefaciens GV3101 (i.e. agrobacterium is suspended in an induction medium) [¶426]. The explants were blotted dry and cocultivated on MS20 medium with 40 μg.I-1 acetosyringone and 10 μM β-estradiol for stable transformation of the plant cell. Scheres teaches that the induction medium with estradiol for tomato explants transformed with the construct resulted in higher shoot formation than conventional shoot induction medium with plant growth regulators (NAA+BAP and zeatin) (i.e. wherein the induction medium is selected from the group consisting of callus, shoot, or root induction medium) [¶429].
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claim 12 is rejected under 35 U.S.C. 103 as being unpatentable over Scheres as applied to claims 11 and 13-15 above, and further in view of Maher et al., Plant gene editing through de novo induction of meristems, Nature Biotechnology, 2020, 38:84-89 (as cited in IDS filed 07/18/2024).
Claim 12 recites the method of claim 11, wherein the plant is an adult plant.
The method of claim 11 is anticipated by Scheres, as rejected above. Scheres does not explicitly teach the method of transforming a plant with a PLT construct wherein the plant is an adult plant. However, Maher teaches an exogenous morphogenic regulator-based in vivo transformation method on soil-grown plants which avoids tissue culture entirely. DRs from Arabidopsis thaliana, such as WUSHEL (WUS), BABY BOOM (BBM) and SHOOT MERISTEMLESS (STM) are delivered with Agrobacterium to somatic cells of whole plants to induce the formation of de novo meristems [pg. 1, col. 1, ¶1]. Maher teaches that the expression of DRs creates transgenic and gene-edited shoots through de novo meristem induction, which are able to produce flowers and seeds, ultimately transmitting transgenes and gene edits to the next generation [pg. 1, col. 2, ¶1].
Maher teaches that the Agrobacterium cultures were grown overnight in growth medium and suspended in infiltration medium (10 mM MES, 150 μM acetosyringone, 10 mM MgCl2). N. benthamiana plants were grown to maturity (63-66 d) (i.e. adult plants), visible meristems were removed, and the plants immediately inoculated with A. tumefaciens cultures at the wound sites using syringes [pg. 90, col. 1, ¶6]. Potato, tomato, and grape plants were propagated asexually on growth medium before exiting meristems were removed and the wound inoculated as in N. benthamiana [pg. 90, col. 1, ¶2 & 3]. Maher teaches that transgenic shoots on tobacco, tomato, potato and grape can be developed in a fraction of the time required for traditional tissue culture methods [pg. 88, col. 2, ¶3].
Scheres teaches a method of transforming a plant with a PLT construct and teaches that PLT is one of several important developmental regulator proteins. Scheres teaches that PLT proteins accumulate in the quiescent center to form a gradient of auxin, the plant hormone important to the establishment of shoot and root apical meristems [¶97]. PLT proteins can regulate PIN gene family expression, a family that plays a significant role in auxin transport and distribution, suggesting a positive feedback loop to maintain high auxin and PLT levels in the stem cell niche (the region where stem cells are located and maintained) [¶97]. Based on the method of DR transformation into in vivo plants as taught by Maher, and the importance of PLT proteins to plant development, particularly to shoot and root meristematic growth, and the PLT construct taught by Scheres, it would have been prima facie obvious to one of ordinary skill in the art at the time of filing to modify the method of DR transformation into in vivo plants as taught by Maher with the introduction or increased expression of a PLT protein of Scheres to overexpress this developmental regulator gene to enhance meristematic growth. One would have been motivated to do so as Maher teaches that the method of in vitro injection of the Agrobacterium culture with the transgene is a much more efficient strategy than traditional tissue culture methods. The PLT protein would be an obvious protein to try as Maher already demonstrates the use of other developmental regulator proteins, such as WUS, BBM, and STM.
Conclusion
No claims are allowed.
Contact Information
Any inquiry concerning this communication or earlier communications from the examiner should be directed to EMILY K. JOHNSON whose telephone number is (571)272-5761. The examiner can normally be reached Monday - Friday 7:30 am - 5:00 pm.
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/EMILY K JOHNSON/Examiner, Art Unit 1662
/BRATISLAV STANKOVIC/Supervisory Patent Examiner, Art Units 1661 & 1662