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 .
Claim Status
Claims 1-2, 4, 6, 9, 14-15, 18, 20-21, 23, 25-28, 32, 34-37, 39, and 42 are pending.
Claims 1-2, 4, 6, 9, 14-15, 18, 20-21, 23, 25-28, 32, 34-37, 39, and 42 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 1-2, 4, 6, 9, 14-15, 18, 20-21, 23, 25-28, 32, 34-37, 39, and 42 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.
Claims 1-2, 4, 6, 9, 14-15, 18, 20-21, 23, 28, 32, 35, 39, and 42 are rejected as indefinite for the recitation of “plant LysM receptor polypeptide” and related genus terms including “plant non-NFR1 LysM receptor polypeptide” and “plant non-CERK6 LysM receptor polypeptide”. As described in the specification (paragraph 0005-0006), plant LysM receptors are receptor-like kinase comprising an extracellular domain containing LysM motifs, a transmembrane domain, a juxtamembrane (JM) domain, and an intracellular kinase domain, and include diverse classes such as CERKs and NFRs that mediate distinct signaling pathways.
However, this description does not provide objective boundaries for determining which proteins qualify as plant LysM receptor polypeptides within the meaning of the claims. In particular, the specification describes the term at a high level of generality based on the presence of certain domains and broad functional groupings, but does not set forth sufficiently definite criteria such as the required number, arrangement, or sequence conservation of LysM motifs, the degree of sequence identity or homology requried, whether all recited domains must be present and functional, what receptor activity or signaling capability is required, or how such activity is to be assessed. Thus, neither the claims nor the specification provide an objective standard for determining which proteins fall within the recited genus and which do not.
This ambiguity is especially significant because the term receptor denotes a functional classification, yet the claims do not provide a reasonably certain boundary for distinguishing the recited plant LysM receptor polypeptides from other structural relate LysM-containing or receptor-linked proteins. The uncertainty in genus membership is further compounded because the claims broadly recite modifications to JM zone 4 and, in some claims, optional modifications to kinase regions and extracellular domains, as well as recited signaling outcomes. Without first being able to determine with reasonable certainty what proteins qualify as the required plant LysM receptor polypeptides, one of ordinary skill in the art would not be reasonable apprised of the metes and bounds of the claimed invention.
Dependent claims 25-27, 34, 36, and 37 are included in this rejection because they do not include additional limitations to resolve the ambiguity.
Claims 1, 6, 15, 21, 23, 32, and 39 are rejected as indefinite for the recitation “corresponds to” or “corresponding to” to define the boundaries of the claimed JM zone 4, kinase C-terminus region, and/or kinase N-terminus region by reference to amino acid positions in SEQ ID NO:1 and/or SEQ ID NO:8, but do not set forth the alignment method or parameters by which such correspondence id determined.
For example, claim 1 recites that “the first JM zone 4 with the corresponding amino acids from a second JM zone 4 of a second plant LysM receptor polypeptide, wherein the first JM zone 4, the second JM zone 4, or both correspond to amino acids 305 to 327 when aligned to SEQ ID NO: 1 or correspond to amino acids 303 to 325 when aligned to SEQ ID NO: 8”, and claim 39 similarly recites identifying a region “corresponding to” specified amino acids after aligning a candidate receptor to SEQ ID NO:1 or SEQ ID NO:8. However, the claims do not specify which alignment algorithm is to be used, whether the alignment is global or local, how insertions and deletions are to be treated, what gap penalties apply, or how alternative reasonable alignments are to be resolved. Because different alignment methods or parameters can yield different residue-to-residue mapping, one of ordinary skill in the art would not be able to demine with reasonable certainty which residues of candidate receptor constitute the claimed region. Accordingly, the metes and bounds of the claims are not reasonably certain.
Claims 1, 2, 6, 14, 23, 28, 32, and 39 are rejected as indefinite because the claims require substitution of amino acids in a first region with “the corresponding amino acids” or “corresponding amino acid residues” from a second region, but fail to define what makes one amino acid in the second region the “corresponding” amino acid for an amino acid in the first region.
The claims do not make clear whether “corresponding” refers to the same numerical position, the same aligned position, the same homologous position, or some other positional or functional relationship. This ambiguity is especially significant because certain claims, such as claim1, also permit insertion and deletion, such that the first and second regions may differ in length and may not have a simple one-to-one residue relationship. As a result, one of ordinary skill in the art would not be able to determine with reasonable certainty which amino acid(s) from the second region must replace which amino acid(s) in the first region. Therefore, the scope of the claims in not reasonably certain.
Claim 1 is rejected as indefinite for the additional reason that the limitation “wherein the first JM zone 4, the second JM zone 4, or both correspond to amino acids 305 to 327 when aligned to SEQ ID NO: 1 or correspond to amino acids 303 to 325 when aligned to SEQ ID NO: 8” fails to make reasonably certain which JM zone must satisfy the positional limitation.
In particular, it is unclear whether the limitation is met when only the first JM zone 4 corresponds to the recited residues, when only the second JM zone 4 corresponds to the recited residues, or only when both do. It is further unclear whether the same reference sequence and same alignment must be used for both regions. Because the claim does not clearly identify which region or combination of regions must meet the stated positional requirement, the scope of claim 1 is not reasonably certain.
Claim 20 is rejected as indefinite for the recitation of “the first extracellular domain is modified as compared to the amino acid sequence of the corresponding unmodified plant LysM receptor polypeptide”, but fails to identify what protein constitutes the “corresponding unmodified” polypeptide.
Given that the claimed LysM receptor polypeptide may already being modified in the JM zone, kinase region, and /or extracellular domain, and may include sequence material derived from more than one receptor polypeptide, it is unclear whether the “corresponding unmodified” polypeptide refers to the original starting receptor, the receptor contributing the extracellular domain, a naturally occurring ortholog, or some other reference sequence. Because the comparison baseline is not reasonably certain, one of ordinary skill in the art would not be able to determine with reasonable certainty whether the extracellular domain is “modified as compared to” the required reference. Accordingly, the scope of claim 20 is indefinite.
Dependent claims 25-27, 34, 36, and 37 are included in this rejection because they do not include additional limitations to resolve the ambiguity.
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.
Written Descriptions
Claims 1-2, 4, 6, 9, 14-15, 18, 20-21, 23, 25-28, 32, 34-37, 39, and 42 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 Federal Circuit has clarified the application of the written description requirement. The court stated that a written description of an invention "requires a precise definition, such as by structure, formula, [or] chemical name, of the claimed subject matter sufficient to distinguish it from other materials". University of California v. Eli Lilly and Co., 119 F.3d 1559, 1568; 43 USPQ2d 1398, 1406 (Fed. Cir. 1997). The court also concluded that "naming a type of material generally known to exist, in the absence of knowledge as to what that material consists of, is not description of that material". Id. Further, the court held that to adequately describe a claimed genus, Patent Owner must describe a representative number of the species of the claimed genus, and that one of skill in the art should be able to "visualize or recognize the identity of the members of the genus". Id.
Claims 1-2, 4, 6, 9, 14-15, 18, 20-21, 23, 28, 32, 35, 39, and 42 and dependent claims 25-27, 34, 36, and 37 are directed to a modified plant LysM receptor polypeptide. The breath of the claimed genus begins with the recited term “plant LysM receptor polypeptide” itself. As described in the specification (paragraph 0005-0006), plant LysM receptors are receptor-like kinase comprising an extracellular domain containing LysM motifs, a transmembrane domain, a juxtamembrane (JM) domain, and an intracellular kinase domain, and include diverse classes such as CERKs and NFRs that mediate distinct signaling pathways. However, this description dotes not provide identifying structural and functional characteristics sufficient to delimit what proteins qualify as members of the claimed genus of plant LysM receptor polypeptides across its full scope.
In particular, the term “LysM receptor” carries a required functional identity, yet the specification does not establish a common structural principle or objective functional boundary by which one of ordinary skill in the art could determine which proteins are true LysM receptors within the meaning of the claims, as opposed to other structurally related LysM-containing or receptor-line proteins. The specification identifies broad domain architecture and example receptor subclasses, but does not explain what minimum structural features, signaling properties, ligand-recognition characteristics, or other unifying criteria are required for membership in the claimed genus. Thus, the specification odes not reasonably convey possession the full genus anchored by the term “plant LysM receptor polypeptide”.
The claims further broadly recite modification of JM zone 4 by insertion, deletion, or substitution of one or more amino acids using corresponding amino acids form a second LysM receptor, including embodiments involving multiple residue substitutions and functional outcomes such as initiation of NFR1-mediated root nodule symbiosis signaling, ROS signaling, or different signaling. Thus, the claims are not limited to a small set of specifically exemplified receptor or substitution, but instead encompass a broad genus of modified LysM receptor polypeptides defined in part by sequence manipulation of JM zone 4 and by the resulting signaling activity.
The specification provides specific and limited examples of modified LysM receptors, inducing: chimeric receptor constructs involving domain swaps between NFR1 and CERK6 transmembrane JM regions (example 1); identification of JM zone 4 as a determinant of symbiotic signaling (example 1); individual amino acid substitutions within JM zone 4, such as M306, A308, K320 and related residues, and their effects on nodulation (example 3); combinatorial substitutions, such as M306T/A308D/K320T, affecting signaling outcomes (example 4); and limited gain-of-function substitutions showing partial restoration of signaling (examples 5-6).
These disclosures are focused on specific receptor pairs, such as NFR1/CERK6 and related constructs, and on a small number of residues within JM zone 4. The specification therefore provides only a limited set of examples within a much broader claimed genus.
The claims encompass: any plant LysM receptor polypeptide; any second LysM receptor as a source of amino acids for JM zone 4 modification; and numerous possible modifications across JM zone 4, including sing-residue, multi-residue, insertion, deletion, and substitution variants, tied to multiple functional outcomes.
In contrast, the specification discloses only a limited number of specific embodiments, primarily involving NFR1/CERK6-based constructs and a small subset of identified JM zone 4 residues. The specification does not provide representative species across the full scope of plant LysM receptors, across the breadth of receptor subclasses, or across the full range of possible JM zone 4 modifications now claimed. Thus, the specification does not demonstrate possession of the full claimed genus.
Although the claims are directed to a broad genus of modified plant LysM receptor polypeptides, neither the claims nor the specification identify a set of common structural features, conserved sequence characteristics, or other unifying structural rules sufficient to define which JM zone 4 modifications across the genus will yield the claimed signaling outcomes. Plant LysM receptors comprise a structurally related but functionally diverse class of receptor-like kinases, including symbiotic receptors (e.g., NFR1/NFR5) and immune receptors (e. g., CERK-type receptors), which mediate distinct signaling pathways (e.g., Cyril Zipfel et. al., NATURE (2017|) VOL 543 pp328-336) (page 331-332, Fig. 3, The formation of ligand-induced receptor kinase complexes and the role of regulatory receptor kinases). Antolín-Llovera (Meritxell Antolín-Llovera et. al., New Phytologist (2014) 204: 791–80) (Abstract, and page 796 left column) and Bigeard (Jean Bigeard et. al., Molecular Plant (2015) 8, 521–539) (page 528, left column paragraph 2) further demonstrates that receptor signaling depends on domain context and specific residue interactions, and that domain swaps or mutations may alter, abolish, or fail to confer signaling.
While the specification identifies certain residues as important in the particular context of the exemplified receptors, such as M306, A308, and K320, it does not establish a general structural principle applicable across: different LysM receptor families; different plant species; different receptor sequence backgrounds; or different combinations of substitutions, insertions, or deletion with JM zone4.
Accordingly, specification does not reasonably convey possession of the full genus by disclosure of either representative species or common structural characteristics correlated with the claimed function.
The claimed invention relies on the premise that modifying JM zone 4 of a plant LysM receptor using amino acids from another LysM receptor can alter receptor signaling, including conferring NFR1-meidated symbiosis signaling, ROS signaling, or other signaling outcomes. However, prior art and the specification demonstrate that the relationship between JM zone 4 sequence and signaling function is highly specific and context-dependent (e.g., Antolin-Llovera, Abstract, and page 796 left column).
For example, the disclosure indicate that: substitution of certain individual residues, such as M306 or A308, can impair nodulation, whereas other changes do not (paragraph 0195); certain combination of substitutions are required to obtain functional signaling (paragraph 0200); and some constructs fail to restore or confer the desired function (paragraph 0272). This further demonstrates that receptor signaling depends on precise domain context and residue interactions, and that domain swaps or mutations may alter, abolish, or fail to confer signaling.
These results show that signaling outcomes are highly sensitive to particular residues and combinations of residues, and that the effect of modification is not predictable form the disclosure as a general matter. Despite this , the claims broadly cover modification throughout JM zone 4 across a broad genus of LysM receptors, without being limited to the specific receptors, residues, or substitution patterns actually described. As such, the specification does not reasonably possession of the full scope of the claimed invention.
Claim Rejections - 35 USC § 102
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 1-2, 4, 6, 9, 14-15, 18, 20-21, 23, 25-27, 32, 34-37, 39, and 42 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Bozsoki (Zoltan Bozsoki et. al., Science (2020) 369, 663–670).
Under the broadest reasonable interpretation, claim 1 recites a modified plant LysM receptor polypeptide in which a first JM zone 4 is modified by insertion, deletion, or substitution of one or more amino acids in the first JM zone 4 with corresponding amino acids from a second JM zone 4 of a second plant LysM receptor polypeptide. The recitation “one or more amino acids” encompasses substitution of multiple amino acids, including substitution of the entire corresponding JM zone 4 region, because the claim does not limit the modification to only a subset of residues or exclude complete replacement of the corresponding aligned region.
Bozsoki discloses plant LysM receptor polypeptides including NFR1 and CERK6 (Fig. S1, B, also see below), each comprising extracellular LysM domains, a juxtamembrane region (TJ), and a kinase domain (KD) (Fig 1A). Bozsoki further identify the TJ region as a discrete contiguous region positioned between the ectodomain and kinase domain, shown for NFR1 between residues A226 and L327 and for CERK6 between residues G226 and L325 (Fig 1A, also see alignment below). Furthermore, the sequence alignment demonstrates that SEQ ID NO:1 corresponds to LjNFR1 and SEQ ID NO:8 corresponds to LjCERK6 (see alignment).
Bozsoki additionally discloses chimeric LysM receptor polypeptides in which the TJ
region of one receptor is replaced with the corresponding TJ region from the other receptor, as shown by the domain-swap constructs in Fig. 1D (e.g., construct 13-16). Thus, Bozsoki expressly disclose a first LysM receptor polypeptide having its corresponding JM/TJ region modified by substitution of amino acids in that region with corresponding amino acids from the TJ region of a second plant LysM receptor polypeptide.
Because replacement of the entire corresponding TJ/JM region necessarily includes substitution of one or more amino acids in the first JM zone 4 (juxtamembrane is zone 4, from instant application paragraph 0004) with corresponding amino acids from the second JM zone 4, and because the swapped regions are corresponding aligned regions of the respective plant LysM receptor polypeptides, Bozsoki disclose each and every element of claim 1 required by the claim. Therefore, claim 1 is anticipated by Bozsoki.
Under BRI, the limitations of claim 2(i), 2(ii), and 2(iii) are joined by “and/or”, such that satisfaction of any one of these alternatives is sufficient to meet the claim.
Claim 2 recites the modified plant LysM receptor polypeptide of claim 1, wherein: (i) the first JM zone 4 is modified by substituting at least two or at least three amino acid residues in the first JM zone 4 with corresponding amino acid residues that are different in the second JM zone 4.
Bozsoki discloses modification of the first JM/TJ region by substitution of multiple corresponding amino acid residues from a second plant LysM receptor polypeptide (see below alignment of the fragments). In particular, the disclosed swap results in substitution of six amino acid residues in the corresponding JM zone 4 region, which satisfies claim 2(1)’s requirement of substituting at least two or at least three amino acid residues in the first JM zone 4 with corresponding amino acid residues that are different in the second JM zone 4.
Therefore, claim 2 is anticipated by Bozsoki.
Under the broadest reasonable interpretation, claim 4 recites modified plant LysM receptor polypeptide of claim 2. Claim 4 is satisfied by any one of subparts (i)-(iv). Applicant’s claim 4(i) requires that the second JM zone 4 is able to imitate NFR1-mediated root nodule symbiosis signaling.
Bozsoki discloses NFR1 as a plant LysM receptor involved in root nodule symbiosis signaling and further discloses use of the corresponding NFR1-derived TJ/JM region as the donor region in domain-swap chimeric receptors (page 664 Fig. 1A; Fig. 1D). Bozsoki discloses “Intact NFR1 (1 in Fig. 1, B and C) and all chimeras with the ectodomain of NFR1 (2, 3, and 4 in Fig. 1B) induced nodule formation on nfr1-1 (nfr1) mutants” (page 663, column 3). Accordingly, the second JM zone 4 in Bozsoki, when derived from NFR1, is able to initiate NFR1-mediated root nodule symbiosis signaling, as required by claim 4(i).
Therefore, Bozsoki discloses each and every element of claim 4, arranged as required by the claim, and claim 4 is anticipated.
Claim 6 is interpreted under the broadest reasonable interpretation, is directed to a modified plant non-NFR1 LysM receptor polypeptide engineered for NFR1-mediated root nodule symbiosis signaling, wherein the receptor comprised either: (i) a first JM zone 4 modified by substitution of one or more amino acids in the first JM zone 4 with corresponding amino acids from a second JM zone 4 from an NFR1 LysM receptor polypeptide with NFR1-mediated root nodule symbiosis signaling. Because claim 6 uses “or” between embodiments (1) and (2), and “and/or” between subparts (i) and (ii) of embodiment (1), the claim is met if the reference discloses embodiment (1)(i) alone.
Claim 6(1)(i) is interpreted as encompassing a modified plant non-NFR1 LysM receptor polypeptide in which the first JM zone 4 is modified by substitution of one or more amino acids in the first JM zone 4 is modified by substitution of one or more amino acids in the first JM zone 4 with the corresponding amino acids from a second JM zone 4 from an NFR1 LysM receptor polypeptide with NFR1-mediated root nodule symbiosis signaling. Because the claim recites substitution of “one or more amino acids”, the limitation reads not only on substitution of a single amino acid, but also on substitution of multiple amino acids or the entire corresponding JM/TJ region. Therefore, a disclosed Swap of the corresponding JM/TJ region between a non-NFR receptor and an NFR1 receptor falls within the scope of claim 6(1)(i).
Bozsoki discloses chimeric plant LysM receptor polypeptides constructed from NFR1 and CERK6 domains, including receptors comprising extracellular LysM domains, a juxtamembrane region designated TJ, and a kinase domain (Fig. 1A). Bozsoki further discloses that the TJ region is a discrete intracellular region between the ectodomain and kinase domain and corresponds to the claimed JM zone 4. Bozsoki further discloses domain-swap chimeras in which the corresponding TJ/JM region of one receptor is substituted with the corresponding TJ/JM region form the other receptor (Fig. 1D)
Bozsoki further discloses modified non-NFR1 receptors, including CERK6-based chimeric receptors carrying NFR1-derived regions, that were engineered and evaluated for NFR1-mediated root nodule symbiosis signaling in nfr1-1 mutant plants (page 663, column 3). Bozsoki further discloses chimeric LysM receptors that are not intact NFR1 receptors and that were designed and tested for restoration of nodulation in nfr1-1 mutants, thereby demonstrating modified non-NFR1 chimeric receptors engineered for NFR1-mediated root nodule symbiosis signaling (Fig. 1B and page 663, column 3).
Accordingly, Bozsoki discloses a modified plant non-NFR1 LysM receptor polypeptide engineered for NFR1-mediated root nodule symbiosis signaling, comprising a first JM zone 4 modified by substitution of one or more amino acids in the first JM zone 4 with corresponding amino acids from a second JM zone 4 from an NFR1 LysM receptor polypeptide, as required by claim 6(1)(i). Therefore, Bozsoki discloses each and every element of claim 6, arranged as required by the claim.
Claim 9 is interpreted under the broadest reasonable interpretation as encompassing a modified plant LysM receptor polypeptide of claim 6 in which: (1) the modified plant LysM receptor polypeptide is the non-NFR1 LysM receptor polypeptide, and wherein the first kinase C-terminus region is modified by substituting one or more amino acids of T467, S471, S473, .. or G621 of SEQ ID NO: 8 with one or more amino acids of I469, N473, T475, … or V622 of SEQ ID NO: 1.
Under BRI, the recitation of substituting “one or more amino acids” reads not only on substitution of any single listed amino acid, but also on substitution of multiple listed amino acids, including substitution of the entire recited kinase C-terminus region. Thus, the claim encompasses embodiments in which the claimed residues are replaced individually, in subsets, or as part of a larger domain-level substitution.
Bozsoki discloses modified plant LysM receptor polypeptides generated as chimeric receptors between which NFR1 and CERK6, in which intracellular domains, including the transmembrane/juxtamembrane (TJ) region and the kinase domain (KD), are exchanged between the two receptors (See Fig. 1A, Fig. 1B, Fig. 1D). Bozsoki discloses “Chimeras enabled nodulation of nfr1 with different efficiencies”, and that “exchanging both TJ and kinase with CERK6 in 4 had a significant effect on nodulation” (in page 663, column 3).
With respect to claim 9(1), Bozsoki discloses a non-NFR1 LysM receptor polypeptide in which the first kinase C-terminus region is modified by substitution with the corresponding amino acids from an NFR1 LysM receptor polypeptide. In particular, Bozsoki’s kinase-domain swap chimeras replace the CERK6 kinase-domain sequence with the corresponding NFR1 kinase-domain sequence. Because the claim requires substitution of “one or more amino acids” in the recited first kinase C-terminus region, Bozsoki’s disclosed domain swap reads on the claim under the broadest reasonable interpretation, since replacement of the entire corresponding kinase-domain region necessarily includes substitution of at least one, and indeed multiple, amino acids within the claimed set of residues.
Sequence alignment between CERK6 and NFR1 further shows that the residues recited in claim 9 correspond positionally between the two receptors, such that swapping the kinase region necessarily substitutes the claim CERK6-aligned residues with the corresponding NFR1-aligned residues, as required by claim 9(1). For example, at positions corresponding to S541 and Q542 of SEQ ID NO:8, the aligned NFR1 residues are N542 and K543 of SEQ ID NO:1; thus, a kinase-region swap would replace the NFR1 residues NK with the CERK6 residues SQ at those corresponding positions, demonstrating that the recited amino-acid “substitutions” result from the disclosed domain swap. The same reasoning applies to the other specifically recited residues within the claimed kinase C-terminus region.
Therefore, Bozsoki expressly or inherently discloses “the modified plant LysM receptor polypeptide is the non-NFR1 LysM receptor polypeptide, and wherein the first kinase C-terminus region is modified by substituting one or more amino acids of (the recited residues) of SEQ ID NO: 8 with one or more amino acids of (the recited residues) of SEQ ID NO: 1.
Accordingly, claim 9 is anticipated by Bozsoki.
Claim 14 is interpreted under the broadest reasonable interpretation as requires the modified plant LysM receptor polypeptide of claim 1 to further comprise: (1) a first kinase N-terminus region modified by substitution of one or more amino acids with the corresponding amino acids from a second kinase D-terminus region from an NFR1 LysM receptor polypeptide
Claim 15 is interpreted under the broadest reasonable interpretation as narrowing claim 14 by reciting, with respect to the kinase N-terminus region, the specified aligned residues of SEQ ID NO: 1 and/or that the first kinase N-terminus region is modified by substituting one or more of those residues with the corresponding recited residues of SEQ ID NO: 8. Because the limitations in claim 15 are joined by “and/or”. The ROS-signaling clause is not required for every embodiment, and the claim is met by the recited structural kinas N-terminus limitations alone.
As discussed with respect to claim 1, Bozsoki disclosed modified plant LysM receptor polypeptides generated as chimeric receptors between NFR1 and CERK6, Bozsoki further discloses exchange of intracellular regions, including the TJ and kinase domain regions, between that two receptors (Gig. 1a, Gig. 1D). such exchange discloses a modified plant LysM receptor polypeptide further comprising a first kinase N-terminus region modified by substitution of one or more amino acids with the corresponding amino acids from a second kinase N-terminus region from a CERK6 LysM receptor polypeptide, as recited in claim 14.
With respect to claim 15, the alignment between NFR1 and CERK6 shows the corresponding amino acid positions in the kinase N-terminus region, and Bozsoki’s disclosed kinase-region swap therefore discloses the first kinase N-terminus region corresponding to the recited residues and discloses substitution of one or more of the residues of SEQ ID NO:1 with the corresponding recited residues of SEQ ID NO: 8 (see alignment and marked residues below). Because claim 15 recites these structural limitations in the alternative through “and/or”, disclosure of the recited structural kinase N-terminus limitations is sufficient.
Therefore, Bozsoki anticipates the claim 14 and 15.
Claim 18 is interpreted under the broadest reasonable interpretation as encompassing the modified plant LysM receptor polypeptide of claim 6(1)(i), claim 18(1) requires a modified plant non-NFR1 LysM receptor polypeptide comprising a first JM zone 4 modified by substitution of one or more amino acids corresponding amino acids from the second kinase C-terminus region from a second JM zone 4 from an NFR1 LysM receptor polypeptide, and further comprising a first kinase C-terminus region modified by substitution of one or more amino acids with corresponding amino acids from a second kinase C-terminus region from the NFR1 LysM receptor polypeptide, wherein the modified plant non-NFR1 LysM receptor polypeptide is able to initiate NFR1- mediated root nodule symbiosis signaling. Because the claim is written in the alternative, it is sufficient that the prior art discloses this first recited embodiment.
As set forth with respect to claim 6(1)(i), Bozsoki discloses modified plant LysM receptor polypeptides generated as chimeric receptors between Lotus japonicus NFR1 and CERK6, including receptors in which intracellular regions, including the transmembrane /juxtamembrane (TJ) region and kinase domain, are exchanged between the two receptors (Gig. 1A, Fig. 1B). Bozsoki therefore discloses that chimera 4 contains exchange of both the TJ and Kinase regions and that this construct formed nodule in nfr1-1mutants (Gig. 1B), and “whereas exchanging both TJ and kinase with CERK6 in 4 had a significant effect on nodulation. Only 2 out of the 33 plants expressing 4 formed nodules (Fig. 1B)” (page 663, column 3).
Thus, Bozsoki discloses a modified non-NFR1 LysM receptor polypeptide having the intracellular region modified by substitution with the corresponding intracellular region from NFR1, including the first kinase C-terminus region modified by substitution of one or more amino acids with the corresponding amino acids from the second kinase C-terminus region from the NFR1 LysM receptor polypeptide. Because the claim requires substitution of only one or more amino acids, the disclosed exchange of the kinase region necessarily encompasses substitution of at least one, and indeed multiple, corresponding amino acids withing he claimed kinase C-terminus region. Bozsoki also expressly discloses that chimera 4 is able to initiate NFR1-mediated root nodule symbiosis signaling, as evidenced by nodule formation in nfr1-1 mutants.
Accordingly, claim 18 is anticipated by Bozsoki.
Claim 20 is interpreted under the broadest reasonable interpretation as requiring the modified plant LysM receptor polypeptide of claim 1 to further comprise a first extracellular domain, wherein the first extracellular domain is modified as compared to the amino acid sequence of the corresponding unmodified plant LysM receptor polypeptide.
As set forth disclosed from claim 1, Bozsoki discloses modified plant LysM receptor polypeptides formed as chimeric receptors between NFR1 and CERK6, wherein receptor domains including the ectodomain, TJ region, and kinase domain are exchanged (Fig. 1A). since the ectodomain is the extracellular domain of the receptor, Bozsoki discloses a modified plant LysM receptor polypeptide of claim 1 that further comprises a first extracellular domain, wherein the first extracellular domain is modified relative to the amino acid sequence of the corresponding unmodified plant LysM receptor polypeptide, as recited in claim 20. Therefore, claim 20 is anticipated.
Under BRI, claim 21 requires a genetically modified plant or part thereof comprising a modified plant LysM receptor polypeptide in which a CERK6-type JM zone 4 (e.g., SEQ ID NO:13) is modified by insertion, deletion, or substitution with corresponding amino acids from an NFR1-type JM zone 4 (e.gl, SEQ ID NO:6). Wherein the modified plant LysM receptor polypeptides is able to initiate NFR1-mediated root nodule symbiosis signaling.
Bozsoki discloses, SEQ ID NO 6 (IMVAKSMEFS YQELAKATNN FSL) is same as SEQ ID NO 1 (305 to 327: IMVAKS MEFSYQELAK ATNNFSL) which is from LjCERF6; SEQ ID NO:13 (ITVDKSVEFS YDELATATDN FSL) is same as SEQ ID NO:8 (303-325: ITVDKSVE FSYDELATAT DNFSL) which is from LjNFR1 (Supplementary, Fig. S1).
Set forth above with respect to claim 1, Bozsoki discloses chimeric plant LysM receptor polypeptides between NFR1 and CERK6, including constructs in which the TJ (JM zone 4) region is exchanged (Fig. 1A), which refers to the replacement of SEQ ID NO: 13 with SEQ ID NO:6. Bozsoki further discloses “Intact NFR1 (1 in Fig. 1, B and C) and all chimeras with the ectodomain of NFR1 (2, 3, and 4 in Fig. 1B) induced nodule formation on nfr1-1 (nfr1) mutants”(page 663, column 3), thereby evidencing that the modified receptor is able to initiate NFR1-mediated root nodule symbiosis signaling in plant. Because the domain exchange replaces corresponding amino acids between CERK6 and NFR1, Bozsoki discloses the claimed JM zone 4 modification.
Under the broadest reasonable interpretation, Claim 23 further requires that the modified plant LysM receptor polypeptide is a modified non-NFR1 LysM receptor polypeptide, and that the first JM zone 4 is modified by substitution of one or more amino acids with corresponding amino acids from a second JM zone 4 from an NFR1 LysM receptor polypeptide with NFR1-mediated root nodule symbiosis signaling.
Under the broadest reasonable interpretation, Claim 25 further requires that the plant of claim 21, wherein the plant is selected from the group consisting of cassava, yam, sweet potato, corn, cowpea, rice, barley, wheat, Trema spp., apple, pear, plum, apricot, peach, almond, walnut, strawberry, raspberry, blackberry, red currant, black currant, melon, cucumber, pumpkin, squash, grape, bean, soybean, pea, chickpea, pigeon pea, lentil, Bambara groundnut, lupin, pulses, Medicago spp., Lotus spp., forage legumes, indigo, legume trees, and hemp.
Bozsoki discloses that the genetically modified plant is Lotus japonicus (page 663, Abstract)
Claim 26 recites a method of producing the genetically modified plant or part thereof of claim 21, comprising introducing a genetic alteration to the plant comprising a first nucleic acid sequence encoding the modified plant LysM receptor polypeptide or the modified plant non-NFR1 LysM receptor polypeptide.
Claim 27 recites the method of claim 26, wherein the nucleic acid sequence is operably linked to a promoter, wherein the promoter is a root specific promoter, an inducible promoter, a constitutive promoter, or a combination thereof.
Bozsoki expressly discloses the construct is under ubiquitin promoter (Generation of plant expression vectors (Fig. S1)).
Accordingly, claims 21, 23, 25-27 are anticipated by Bozsoki.
Claim 32 is interpreted under the broadest reasonable interpretation, claim 32 requires an expression vector comprising a nucleic acid encoding a modified plant LysM receptor polypeptide operable linked to at least one expression control sequence, wherein the polypeptide comprises a modified JM zone 4.
Bozsoki discloses generation of plant expression vectors comprising nucleic acids encoding chimeric plant LysM receptor polypeptides between NFR1 and CERK6 (Fig. S1). Specifically, Bozsoki discloses that plasmids constraining gene fragments encoding the respective domains of LjNFR1 and LjCERK6 were assembled and cloned into plant expression vectors, and that Nfr1, Cerk6, or chimeric genomic sequences were cloned downstream of the 35S promoter in a binary vector backbone (Generation of plant expression vectors (Fig. S1)).
Thus, Bozsoki discloses an expression vector, isolated DNA molecule, or recombinant nucleic acid comprising a nucleic acid encoding a modified plant LysM receptor polypeptide operably linked to an expression control sequence (promoter).
As set forth above with respect to claim 1, Bozsoki further discloses that the chimeric receptor comprises a modified JM zone 4 (TJ region) resulting from exchange between CERK6 and NFR1. Because the chimeric constructs encode these modified receptors, the disclosed expression vector necessarily comprise nucleic acids encoding a modified plant LysM receptor polypeptide having a modified JM zone 4, as recited.
Claim 34 recites a bacterial cell or an Agrobacterium cell comprising the expression vector, isolated DNA molecule, or recombinant nucleic acid of claim 32.
Bozsoki discloses that Agrobacterium rhizogenes strain AR1193 and Agrobacterium tumefaciens strain AGL1 are used for transformation experiments, and that these Agrobacterium strain carried the gene of interest for plant transformation (Supplementary, Materials and Methods).
Claim 35 recites genetically modified plant, plant part, plant cell, or seed comprising the expression vector, isolated DNA molecule, or recombinant nucleic acid of claim 32.
Claim 36 recites a composition or kit comprising the expression vector, isolated DNA molecule, or recombinant nucleic acid of claim 32 with a bacterial cell or Agrobacterium cell.
Claim 37 recites a method of initiating NFR1-mediated root nodule symbiosis signaling comprising: introducing a genetic alteration via the expression vector, isolated DNA molecule, or recombinant nucleic acid of claim 32 to a cell.
As set forth above with respect to claim32, Bozsoki further discloses introducing these constructs into plant cells via Agrobacterium rhizogenes-mediated hairy toot transformation, wherein A. rhizogenes carrying the gene of interest is applied to plant tissue to generated transformed root cells (Materials and Methods, “Hairy root transformation” in Supplementary).
Bozsoki additionally discloses that the resulting transformed plant material induces nodule formation in nfr1-f mutants (Fig. 1B), thereby demonstrating that the introduced construct initiates NFR1-mediated root nodule symbiosis signaling.
Accordingly, claims 32, 34-37 are anticipated by Bozsoki.
Under BRI, claim 39 recites a method generating a modified plant LysM receptor polypeptide, comprising: (a) aligning an amino acid sequence of a candidate receptor to SEQ ID NO: 8 to identify a first JM zone 4; (b) modifying the first JM zone 4 by substituting at least two or at least three amino acid residues in the first JM zone 4 with corresponding amino acid residues that are different in a second JM zone 4; and (c) generating the modified plant LysM receptor polypeptide.
Bozsoki discloses a method of generating modified plant LysM receptor polypeptides by producing chimeric receptors between LiNFR1 and LjCERK6, which correspond to the same receptor sequences as SEQ ID NO: 1 and SEQ ID NO: 8, respectively (see sequences and alignment below). Bozsoki further discloses exchange of the TJ region (JM zone 4) between these receptors, as shown in Fig 1A and the supplementary sequence information.
Because Bozsoki uses the same receptor sequences and the same corresponding TJ/JM region boundaries, Bozsoki discloses, aligning the amino acid sequence of the candidate receptor to the corresponding reference receptor sequence in order to identify the first JM sone 4 by substituting the corresponding amino acids from the second JM zone4 through domain exchange between NFR1 and CERK6. Since the disclosed swap replaces the region with corresponding sequence from the second receptor, the prior art necessarily substitutes at least two or at least three amino acid residues that differ between the two JM zone 4 sequences. Bozsoki then generated the resulting modified plant LysM receptor polypeptide as a chimeric receptor.
Claim 42 recites the modified plant LysM receptor polypeptide produced by the method of claim 39.
Claims 39 and 42 are anticipated by Bozsoki.
[AltContent: rect][AltContent: rect][AltContent: rect][AltContent: rect]
PNG
media_image1.png
448
712
media_image1.png
Greyscale
[AltContent: textbox (LjNFR1(305-327))][AltContent: textbox (LjCERK6 (303-325))]
PNG
media_image2.png
288
699
media_image2.png
Greyscale
[AltContent: rect]
PNG
media_image3.png
878
1770
media_image3.png
Greyscale
PNG
media_image4.png
384
1772
media_image4.png
Greyscale
[AltContent: rect]
PNG
media_image5.png
805
1646
media_image5.png
Greyscale
[AltContent: rect]
PNG
media_image6.png
403
975
media_image6.png
Greyscale
PNG
media_image7.png
572
1694
media_image7.png
Greyscale
Claim Rejections - 35 USC § 103
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 28 is rejected under 35 U.S.C. §103 as being unpatentable over Bozsoki (2020) as applied on claim 21, and in view of Jain (Divya Jain et. al., Current Opinion in Plant Biology (2023, Available online 17 December 2022) 71 pp1-8).
Claim 21 as the teachings of Bozsoki is discussed above.
Claim 28 is interpreted as dependent of claim 21.
Under BRI, claim 28 further requires a method of producing the genetically modified plant or part thereof of claim 21, comprising genetically modifying the plant or part thereof by transforming the plant or part thereof with one or more gene editing components that target an endogenous nuclear genome sequence encoding an endogenous plant LysM receptor polypeptide or plant non-NFR1 LysM receptor polypeptide to genetically modify a first JM zone 4 by insertion, deletion, or substitution of one or more amino acids in the first JM zone 4 with the corresponding amino acids from a second JM zone 4 of a second plant LysM receptor polypeptide.
As discussed above with respect to claim 21, Bozsoki teaches modified plant LysM receptor polypeptides generated as chimeric rectors between NFR1 and CERK6, including constructs in which the TJ/JM zone 4 region is exchanged, and further teaches that such modified receptors, when expressed in plant material, are able to initiated NFR1-mediated root nodule symbiosis signaling (page 664, Fig. 1).
Bozsoki does not expressly teach producing the modified plant by transforming the plant with one or more gene editing components targeting an endogenous nuclear genome sequence encoding the receptor.
Jain teaches that gene editing is the process of optimizing endogenous gene function by precise modification of genomic sequences and that cRISPR-Cas9 is a preferred plant genome-editing technology for targeted insertion-deletion mutagenesis (page 1, Abstract). Jain further teaches that gene-editing pipelines for legumes, including Lotus japonicus and Medicago trucatula, had been successfully developed using plant transformation methods (page 2, left column, paragraph 2), and that gene editing was used to study and modify root nodule symbiosis bene, including nodulation receptors and related signaling genes (page 3, figure 1). Jain also teaches that in legumes such as Arachis hypogea, CRISPR-Cas protocols are developed to target NFR1/NFR5 receptor genes, and that edited lines displaced nodulation phenotypes, evidencing the known use of gene-editing components to alter endogenous symbiosis-related receptor genes (page 4, right column, paragraph 1).
It would have been obvious to one of ordinary skill in the art at the time of the invention to implement the receptor modification taught by Bozsoki using the know endogenous gene-editing approaches taught by Jain to transform the plant with one or more gene-editing components targeting the endogenous LysM receptor coding sequence in order to introduce the designed JM zone 4 modification, because Jain teaches that precise genome editing of endogenous symbiosis-related genes in legumes is a known and effective alternative to transgenic introduction for altering nodulation and symbiosis phenotype. One of ordinary skill in the art would have had reason to use such known editing tools to install the Bozsoki receptor modification directly in the endogenous gene to obtain the predictable benefit or producing a plant with the desired nodulation-related receptor function.
Accordingly, it would have been obvious to produce the genetically modified plant of claim 21 by transforming the plant or part thereof with one or more gene editing components targeting an endogenous nuclear genome sequence encoding the receptor, to modify the first JM zone 4 by insertion, deletion, or substitution with corresponding amino acids for a second JM zone 4, as recited in claim 28. Therefore, claim 28 is unpatentable over Bozsoki, in views of Jain.
Conclusion
No claims are allowed.
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.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
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.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/YANXIN SHEN/Examiner, Art Unit 1663
/WEIHUA FAN/Primary Examiner, Art Unit 1663
Prosecution Insights