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’s election without traverse of Group I, claims 1-16 and 18, in the reply filed on December 18th, 2025 is acknowledged.
Claims 17 and 19 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to nonelected inventions, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on December 18th, 2025.
Information Disclosure Statement
The information disclosure statement (IDS) submitted on November 21st, 2024 is acknowledged. The submission is in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statement has been considered by the examiner.
The listing of references in the specification is not a proper information disclosure statement. 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be incorporated into the specification but must be submitted in a separate paper." Therefore, unless the references have been cited by the examiner on form PTO-892, they have not been considered.
Claim Summary
Claims 15 and 16 have been amended. Claims 1-19 are pending. Claims 17 and 19 are withdrawn from consideration as being drawn to a non-elected invention/species. Claims 1-16 and 18 are under examination and discussed in this Office action.
Specification
The use of the terms such as Alexa Fluor, EpiMark, and COVARIS, which are a trade names or a marks used in commerce, have been noted in this application. The terms should be accompanied by the generic terminology; furthermore the terms should be capitalized wherever it appears or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the term.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Claim Objections
Claim 18 is objected to because of the following informalities:
Claim 18 is missing a semi-colon at the end of the recitation “a component for contacting a first portion of the fragmented nucleic acid with a methyl binding protein”. This should be added.
Appropriate correction is required.
Claim Interpretation
Claim 1 recites the limitation “isolating total nucleic acid from the sample”. Turning to the instant specification, “total nucleic acid” in the instant case has been defined as “nucleic acid isolated from a sample that has not been subjected for selection of methylated nucleic acid sequences” (see Page 6, lines 6-7 of the instant specification). Given this definition, reasonable interpretations of total nucleic acid could include genomic DNA isolated from a sample, etc. The provided definition from the instant specification will be applied for interpretation of the phrase “total nucleic acid” throughout this Office Action.
Claim 16 recites the limitation “an expanded FMR1 allele”. Turning to the instant specification, “an expanded FMR1 allele” in the instant case has been defined as “an allele of the FMRI gene that has an expanded CGG triplet repeat region as compared to a normal control. For example, an expanded allele may be classified as a premutation (PM) of about 55-200 CGG repeats, and/or full mutation (FM) of >200 CGG repeats alleles” (see Page 7, lines 1-5 of the instant specification). Given this definition, reasonable interpretations of an expanded FMR1 allele include any detected FMR1 with CGG repeats greater than 55. The provided definition from the instant specification will be applied for interpretation of the phrase “an expanded FMR1 allele” throughout this Office Action.
Claim 18 is drawn to a “system.” The specification recites a “system” wherein the “system” is defined in terms of structural limitations. In addition, the claims recite structural limitations of the “system.” Thus, the “system” is interpreted to encompass any collection of reagents and parts used together that are not necessarily part of a completely integrated single unitary device. Any further interpretation of the word is considered an “intended use” and does not impart any further structural limitation on the claimed subject matter.
Claim Interpretation – 112(f)
The following is a quotation of 35 U.S.C. 112(f):
(f) Element in Claim for a Combination. – An element in a claim for a combination may be expressed as a means or step for performing a specified function without the recital of structure, material, or acts in support thereof, and such claim shall be construed to cover the corresponding structure, material, or acts described in the specification and equivalents thereof.
The following is a quotation of pre-AIA 35 U.S.C. 112, sixth paragraph:
An element in a claim for a combination may be expressed as a means or step for performing a specified function without the recital of structure, material, or acts in support thereof, and such claim shall be construed to cover the corresponding structure, material, or acts described in the specification and equivalents thereof.
The claims in this application are given their broadest reasonable interpretation using the plain meaning of the claim language in light of the specification as it would be understood by one of ordinary skill in the art. The broadest reasonable interpretation of a claim element (also commonly referred to as a claim limitation) is limited by the description in the specification when 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, is invoked.
As explained in MPEP § 2181, subsection I, claim limitations that meet the following three-prong test will be interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph:
(A) the claim limitation uses the term “means” or “step” or a term used as a substitute for “means” that is a generic placeholder (also called a nonce term or a non-structural term having no specific structural meaning) for performing the claimed function;
(B) the term “means” or “step” or the generic placeholder is modified by functional language, typically, but not always linked by the transition word “for” (e.g., “means for”) or another linking word or phrase, such as “configured to” or “so that”; and
(C) the term “means” or “step” or the generic placeholder is not modified by sufficient structure, material, or acts for performing the claimed function.
Use of the word “means” (or “step”) in a claim with functional language creates a rebuttable presumption that the claim limitation is to be treated in accordance with 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph. The presumption that the claim limitation is interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, is rebutted when the claim limitation recites sufficient structure, material, or acts to entirely perform the recited function.
Absence of the word “means” (or “step”) in a claim creates a rebuttable presumption that the claim limitation is not to be treated in accordance with 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph. The presumption that the claim limitation is not interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, is rebutted when the claim limitation recites function without reciting sufficient structure, material or acts to entirely perform the recited function.
Claim limitations in this application that use the word “means” (or “step”) are being interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, except as otherwise indicated in an Office action. Conversely, claim limitations in this application that do not use the word “means” (or “step”) are not being interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, except as otherwise indicated in an Office action.
The claims contain the following limitations:
Claim 18 recites “a component for isolating total nucleic acid from the sample; a component for fragmenting the isolated total nucleic acid to a specific size range; a component for contacting a first portion of the fragmented nucleic acid with a methyl binding protein a component for collecting a fraction of the nucleic acid that is enriched for methylated nucleic acid and a fraction of the nucleic acid that is enriched for unmethylated nucleic acid; a component to perform PCR amplification of the FMRI gene separately for the nucleic acid that is enriched for methylated nucleic acid and the fraction of the nucleic acid that is enriched for unmethylated nucleic acid; and a component to determine if FMRI gene sequences are present in the nucleic acid fraction enriched for methylated nucleic acid and/or the nucleic acid fraction enriched for unmethylated nucleic acid or both”.
Thus, the cited limitations are drawn to generic means plus function. However, this limitation is not being interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, because a review of the specification yields no limiting definition of the generically claimed means.
If applicant intends to have this limitation interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, applicant should indicate where in the specification the specific structure is indicated.
Claim Rejections - 35 USC § 112(b)
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.
Claim 1-16 and 18 are 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 1 recites the limitations "the unbound nucleic acid" and “the bound nucleic acid”. There is insufficient antecedent basis for this limitation in the claim. Neither of these terms have been previously introduced in the claim. Claims 2-16 are also rejected here for their dependence on claim 1 and not further introducing the unbound and bound nucleic acids.
Claim 5 recites the limitation "the total nucleic acid fraction”. There is insufficient antecedent basis for this limitation in the claim. This phrase has not been previously introduced. Claims 1 and 4, from which claim 5 depends, introduces “the total nucleic acid”.
Claim 9 recites the limitation “wherein the at least one primer used to generate an amplification product from the FMR1 gene from the fraction enriched for methylated nucleic acid and/or the fraction enriched for unmethylated nucleic acid and/or the total nucleic acid have the same sequence but are labeled with a different detectable moiety”. It is unclear from this recitation, if the option of having just one primer is exercised in performing the method, how there could be different detectable moieties for a multitude of amplification products. In other words, if there is only one primer, it is not possible for there to be different detectable moieties for each of an amplification product from the FMR1 gene from the fraction enriched for methylated nucleic acid, an amplification product from the FMR1 gene the fraction enriched for unmethylated nucleic acid, and an amplification product from the FMR1 gene the total nucleic acid. There would need to be at least two primers for the option of having the same sequence but different detectable moieties to be possible. This same issue is also noted for claim 11, wherein two labeled primers would have to be used to accomplish the method as claimed. Claim 10 is also rejected here for its dependence on claim 9 and not further clarifying the identified issue. For the purpose of compact prosecution, the recitation of “wherein the at least one primer used to generate an amplification product” is interpreted as requiring at least two primers.
Claim 18 recites the limitation “a component for isolating total nucleic acid from the sample; a component for fragmenting the isolated total nucleic acid to a specific size range; a component for contacting a first portion of the fragmented nucleic acid with a methyl binding protein; a component for collecting a fraction of the nucleic acid that is enriched for methylated nucleic acid and a fraction of the nucleic acid that is enriched for unmethylated nucleic acid; a component to perform PCR amplification of the FMR1 gene separately for the nucleic acid that is enriched for methylated nucleic acid and the fraction of the nucleic acid that is enriched for unmethylated nucleic acid”. There are a few antecedent basis issues in this part of the claim. “The fragmented nucleic acid” in “a component for contacting a first portion of the fragmented nucleic acid with a methyl binding protein” lacks antecedent basis because total nucleic acid was introduced as being fragmented. “The nucleic acid” in “a component for collecting a fraction of the nucleic acid that is enriched for methylated nucleic acid and a fraction of the nucleic acid that is enriched for unmethylated nucleic acid” lacks antecedent basis because it does not acknowledge that the nucleic acid has been fragmented, and also that total nucleic acid was fragmented. Finally, “a component to perform PCR amplification of the FMR1 gene separately for the nucleic acid that is enriched for methylated nucleic acid and the fraction of the nucleic acid that is enriched for unmethylated nucleic acid” lacks antecedent basis because “the nucleic acid that is enriched for methylated nucleic acid” does not acknowledge that the nucleic acid enriched for methylated nucleic acid is a fraction, that the nucleic acid has been fragmented, and also that total nucleic acid was fragmented. Furthermore, “the fraction of the nucleic acid that is enriched for unmethylated nucleic acid” does not acknowledge that the nucleic acid has been fragmented, and also that total nucleic acid was fragmented.
Claim Rejections - 35 USC § 112(a)
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 1-16 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for a human subject, does not reasonably provide enablement for any subject as embraced by the claims. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to use the invention commensurate in scope with these claims.
There are many factors to be considered when determining whether there is sufficient evidence to support a determination that a disclosure does not satisfy the enablement requirement and whether any necessary experimentation is “undue.” See MPEP § 2164. These factors include, but are not limited to: the breadth of the claims, the nature of the invention, the state of the prior art, the level of one of ordinary skill, the level of predictability in the art, the amount of direction provided by the inventor, the existence of working examples, the quantity of experimentation needed to make or use the invention based on the content of the disclosure.
The office has analyzed the specification in direct accordance to the factors outlines in In re Wands. MPEP 2164.04 states: “[W]hile the analysis and conclusion of a lack of enablement are based on factors discussed in MPEP 2164.01(a) and the evidence as whole, it is not necessary to discuss each factor in written enablement rejection.” These factors will be analyzed, in turn, to demonstrate that one of ordinary skill in the art would have had to perform “undue experimentation” to make and/or use the invention and therefore, applicant’s claims are not enabled.
(A) With respect to the breadth of the claims: Claim 1 as currently drafted encompasses a method for detecting FMR1 in methylated and unmethylated fractions of a sample from a subject. “A subject” does not limit the subject to a human subject as described in the specification. Consequently, the breadth of the claims is expansive since they encompass any kind of non-human subject. This can include subjects such as dogs, cats, and birds, among many other examples. Claims 2-16 encompass the same breadth as claim 1 since they do not limit the subject to a human subject.
(B) The nature of the invention: The invention is in the field relating to methods and systems for detecting methylation of the Fragile X FMR1 gene.
(C), (D), (E) With respect to the state of the prior art, the level of one of ordinary skill and predictability of the art: Juppner (Functional properties of the PTH/PTHrP receptor, Bone, August 1995, S39-S42) teaches that despite significant structural conservation, rat, opossum, and human PTH/PTHrP receptor homologs display distinct functional characteristics (Abstract; Pages 39S-40S). This art indicates that there is known functional differences between homologs in different organisms, and therefore inter-species extrapolation would be unpredictable.
The art supports use of specific subjects. However, methods comprising any subject are highly unpredictable.
The invention is drawn to biological molecules, and is therefore in a class of invention which the CAFC has characterized as “the unpredictable arts such as chemistry and biology.” Mycogen Plant Sci., Inc. v. Monsanto Co., 243 F.3d 1316, 1330 (Fed. Cir. 2001). The level of skill in the art is therefore deemed to be high.
(F), (G) With respect to the amount of direction and working examples provided by the applicant: While the Applicant has provided description of a subject including an animal, with specific mammalian examples (Page 5, lines 30-31 to Page 6, lines 1-5), the working examples provided by the Applicant are directed to only human subjects. As noted at Page 30, lines 11-20; Page 31, Table 6; and Page 35, Table 7, the samples used in the working examples are samples from humans. The Applicant has not provided working examples directed towards any other type of subject.
(H) Undue experimentation would be required to practice the invention as claimed due to the amount of experimentation necessary because of the expansive breadth of the claims, the state of the prior art and its high predictability, and the limited amount of guidance in the form of varied working examples in the specification. A skilled artisan recognizes that a subject very broadly refers to any number of different species and thus applicability of the claimed method to a subject as embraced by the claims remains unpredictable, requiring undue experimentation. For example, an artisan would need to test the method and system on an expansive number of different organisms to determine if it is applicable to detecting FMR1 in methylated and unmethylated fractions in said organisms. This reasonably represents undue experimentation.
MPEP §2164.01(a), 4th paragraph, provides that, “A conclusion of lack of enablement means that, based on the evidence regarding each of the above factors, 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 without undue experimentation. In re Wright, 999 F.2d 1157, 1562; 27 USPQ2d 1510, 1513 (Fed. Cir. 1993).
Genentech Inc. v. Novo Nordisk A/S, 42 USPQ2d 1001, 1005 (CA FC), states that, “[p]atent protection is granted in return for an enabling disclosure of an invention, not for vague intimations of general ideas that may or may not be workable,” citing Brenner v. Manson, 383 U.S. 519, 536 (1966) (stating, in the context of the utility requirement, that “a patent is not a hunting license. It is not a reward for search, but compensation for its successful conclusion”). The Genentech decision continued, “tossing out the mere germ of an idea does not constitute enabling disclosure. While every aspect of a generic claim certainly need not have been carried out by an inventor, or exemplified in the specification, reasonable detail must be provided in order to enable members of the public to understand and carry out the invention.” Id. at p. 1005.
After applying the Wands factors and analysis to claims 1-16, in view of the applicant’s entire disclosure, and considering the In re Wright, In re Fisher and Genentech decisions discussed above, it is concluded that the practice of the full scope of the invention as claimed would not be enabled by the written disclosure. Therefore, claims 1-16 are rejected under 35 U.S.C. §112(a) for failing to disclose sufficient information to enable a person of skill in the art to practice the claimed invention to it the full scope embraced by the claims.
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.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1-11 and 14-16 are rejected under 35 U.S.C. 103 as being unpatentable over Latham (US 20120107824 A1), in view of McCarthy (MethylMeter(®): bisulfite-free quantitative and sensitive DNA methylation profiling and mutation detection in FFPE samples, Epigenomics, June 2016, 8, 747-765; cited on the IDS filed November 21st, 2024), as evidenced by Applied Biosciences (DNA Fragment Analysis by Capillary Electrophoresis [online]. Applied Biosciences, [2014] [retrieved on February 13th, 2026]. Retrieved from: https://mugenomicscore.missouri.edu/PDF/fragment-analysis-chemistry-guide.pdf).
Regarding instant claim 1, Latham teaches a method for determining methylation of an FMR1 gene in a sample from a subject comprising the steps of: isolating total nucleic acid from the sample (Pages 10-11, paragraph [0103]), the total nucleic acid comprising both methylated nucleic acid and unmethylated nucleic acid (Page 10, paragraph [0101]; Pages 10-11, paragraph [0103]); contacting a first aliquot of the isolated total nucleic acid with a methylation sensitive enzyme, thereby generating a nucleic acid fraction enriched for methylated nucleic acid (Page 11, paragraph [0105]); conducting PCR amplification of the FMR1 gene using an aliquot of nucleic acid from the fraction enriched for methylated nucleic acid (Page 11, paragraph [0106]); and determining the presence or absence of an FMR1 amplification product in the fraction enriched for methylated nucleic acid (Pages 11-12, paragraphs [0115]-[0119]).
Latham does not teach contacting a first aliquot of the isolated total nucleic acid with a methyl binding protein; isolating a portion of the unbound nucleic acid, thereby generating a nucleic acid fraction enriched for unmethylated nucleic acid; isolating the bound nucleic acid, thereby generating a nucleic acid fraction enriched for methylated nucleic acid; conducting PCR amplification of a target gene using an aliquot of nucleic acid from the fraction enriched for methylated nucleic acid; conducting PCR amplification of a target gene using an aliquot of nucleic acid from the fraction enriched for unmethylated nucleic acid; and determining the presence or absence of a target gene amplification product in both the fraction enriched for methylated nucleic acid and the fraction enriched for unmethylated nucleic acid.
McCarthy, in a reasonably pertinent field, teaches on contacting isolated total nucleic acid with a methyl binding protein (Figure 1; Page 756, column 2, paragraphs 3-4); isolating a portion of the unbound nucleic acid, thereby generating a nucleic acid fraction enriched for unmethylated nucleic acid (Figure 1; Page 756, column 2, paragraphs 3-4); isolating the bound nucleic acid, thereby generating a nucleic acid fraction enriched for methylated nucleic acid (Figure 1; Page 756, column 2, paragraphs 3-4); conducting PCR amplification of a target gene using an aliquot of nucleic acid from the fraction enriched for methylated nucleic acid (Figure 1; Page 756, column 2, paragraphs 3-4); conducting PCR amplification of a target gene using an aliquot of nucleic acid from the fraction enriched for unmethylated nucleic acid (Figure 1; Page 756, column 2, paragraphs 3-4); and determining the presence or absence of a target gene amplification product in both the fraction enriched for methylated nucleic acid and the fraction enriched for unmethylated nucleic acid (Figure 1; Page 756, column 2, paragraphs 3-4).
It would have been obvious to one of ordinary skill in the art, before the effective filing date of the claimed invention, to have modified the method of Latham with the method of McCarthy. Since McCarthy teaches on detecting methylation for target genes, which is reasonably pertinent to the method of Latham, one of ordinary skill in the art would combine the two teachings with a reasonable expectation of success. With regard to the methyl binding protein, one of ordinary skill in the art would have been motivated to make this modification because methyl binding proteins are highly specific for CpG islands and unaffected by the sequencing context of a methylated CpG site (McCarthy, Page 748, column 2, paragraph 2). With regard to also looking at an unmethylated nucleic acid fraction, one of ordinary skill in the art would have been motivated to make this modification because measuring both a methylated and unmethylated fraction allows for determination of methylation percentage (McCarthy, Figure 1). In addition, it would have been obvious to the ordinary artisan that the known techniques of McCarthy could have been applied to the method of Latham with predictable results because the known techniques of McCarthy predictably result in both methylated and unmethylated fractions of input nucleic acid being available for downstream applications, including gene specific PCR (see MPEP 2141(III)). Finally, use of an unmethylated fraction produced by the method of McCarthy, which leaves unmethylated DNA intact, in the method of Latham would amount to simple substitution of one known element for another to obtain predictable results given that the method of Latham is intended to detect intact FMR1 gene (Latham, Page 10, paragraph [0101]); see MPEP 2141(III)).
Regarding instant claim 2, Latham, in view of McCarthy, teaches the method of claim 1. Latham further teaches the method further comprising determining the size of the FMR1 amplification product in a fraction enriched for methylated nucleic acid (Pages 11-12, paragraph [0115]-[0117]). As evidenced by Applied Biosciences, capillary electrophoresis can enable size of an amplification product to be determined via comparison with a size standard (Page 42), such as ROX 1000 (Page 43, size standard table). Latham teaches on using a ROX 1000 ladder (Page 11, paragraph [0115]).
Regarding instant claim 3, Latham, in view of McCarthy, teaches the method of claim 2. Latham further teaches the method further comprising determining the number of CGG repeats at the FMR1 gene for the amplification product in the fraction enriched for methylated nucleic acid (Pages 11-12, paragraphs [0115]-[0118]; Figures 4A-D, bottom graphs; Figures 5A-D, bottom graphs).
Regarding instant claim 4, Latham, in view of McCarthy, teaches the method of claim 1. Latham further teaches the method further comprising performing PCR amplification of the FMR1 gene using a second aliquot of nucleic acid from the total nucleic acid (Page 11, paragraph [0105]) and determining the size of at least one FMR1 amplification product (Pages 11-12, paragraph [0115]-[0117]). As evidenced by Applied Biosciences, capillary electrophoresis can enable size of an amplification product to be determined via comparison with a size standard (Page 42), such as ROX 1000 (Page 43, size standard table). Latham teaches on using a ROX 1000 ladder (Page 11, paragraph [0115]).
Regarding instant claim 5, Latham, in view of McCarthy, teaches the method of claim 4. Latham further teaches the method further comprising determining the number of CGG repeats at the FMR1 gene for the at least one amplification product obtained from the total nucleic acid fraction (Pages 11-12, paragraphs [0115]-[0118]; Figures 4A-D, top graphs; Figures 5A-D, top graphs).
Regarding instant claim 6, Latham, in view of McCarthy, teaches the method of claim 2. Latham does not directly teach wherein determining the size of the FMR1 amplification product in at least one of the fraction enriched for methylated nucleic acid and the fraction enriched for unmethylated nucleic acid comprises comparing the size of the amplification products for the fraction enriched for methylated nucleic acid and/or the fraction enriched for unmethylated nucleic acid to the size of the at least one FMR1 amplification product for total nucleic acid. However, Latham does teach that the total nucleic acid amplification product is mixed with a ROX 1000 ladder as a part of capillary electrophoresis (Page 11, paragraph [0115]). As evidenced by Applied Biosciences, capillary electrophoresis can enable size of an amplification product to be determined via comparison with a size standard (Page 42), such as ROX 1000 (Page 43, size standard table). Therefore, the size of the amplification products of the total nucleic acid would be known. It would then be obvious to compare the amplification products for the fraction enriched for methylated nucleic acid and/or the fraction enriched for unmethylated nucleic acid to the known sizes of the FMR1 amplification product for total nucleic acid to determine the size of the amplification products in other fractions.
Regarding instant claim 7, Latham, in view of McCarthy, teaches the method of claim 6. Latham further teaches wherein the size of the FMR1 amplification product in the fraction enriched for methylated nucleic acid is determined to be the size of the at least one FMR1 amplification product for total nucleic acid (Pages 11-12, paragraphs [0115]-[0118]; Figure 4C; Figure 5B).
Regarding instant claim 8, Latham, in view of McCarthy, teaches the method of claim 1. Latham further teaches wherein at least one primer used to generate an FMR1 amplification product is labeled with a detectable moiety (Page 11, paragraph [0106]).
Regarding instant claim 9, Latham, in view of McCarthy, teaches the method of claim 8. Latham further teaches wherein the at least one primer used to generate an amplification product from the FMR1 gene from the fraction enriched for methylated nucleic acid and the total nucleic acid have the same sequence but are labeled with a different detectable moiety (Page 11, paragraph [0106]).
Regarding instant claim 10, Latham, in view of McCarthy, teaches the method of claim 8. Latham further teaches wherein the detectable moiety is a fluorescent dye (Page 11, paragraph [0106]). Latham earlier teaches that both FAM and HEX are fluorescein dyes (Page 9, paragraph [0089]).
Regarding instant claim 11, Latham, in view of McCarthy, teaches the method of claim 10. Latham further teaches wherein the fluorescent dye used for detection of the amplification products from the total nucleic acid is hexachlorofluorescein (HEX) and the dye used for detection of the amplification product from the fraction enriched for methylated nucleic acid is 6-carboxyfluorescein (FAM) (Page 11, paragraph [0106]), which is the reverse of what is claimed. However, it is taught both are able to be used together (Page 11, paragraph [0106]). It is also taught that the reverse primer they are bound to is the same sequence, and therefore the primers can be used interchangeably on the different fractions (Page 11, paragraph [0106]). Therefore, it would be obvious that they could be used interchangeably for detection of the amplification products of the total nucleic acid and the methylated nucleic acid as this would amount to simple substitution of one known element for another to obtain predictable results (see MPEP 2141(III)).
Regarding instant claim 14, Latham, in view of McCarthy, teaches the method of claim 1. Latham further teaches wherein the size of the FMR1 amplification product in the fraction enriched for methylated nucleic acid is measured by capillary electrophoresis (Pages 11-12, paragraph [0115]-[0117]). As evidenced by Applied Biosciences, capillary electrophoresis can enable size of an amplification product to be determined via comparison with a size standard (Page 42), such as ROX 1000 (Page 43, size standard table). Latham teaches on using a ROX 1000 ladder (Page 11, paragraph [0115]). Latham does not teach that a fraction enriched for unmethylated nucleic acid is measured by capillary electrophoresis. However, given the methods already taught by Latham for the fraction enriched for methylated nucleic acid, as well as the obviousness of an unmethylated fraction in view of McCarthy, it would be obvious to do the same for the fraction enriched for unmethylated nucleic acid because it would amount to simple substitution of one known element for another to obtain predictable results (see MPEP 2141(III)).
Regarding instant claim 15, Latham, in view of McCarthy, teaches the method of claim 1. Latham further teaches the method further comprising determining whether the subject has full methylation of FMR1 (Page 13, paragraph [0132]; Page 16, paragraph [0137]; Table 5: FM=full methylation; Figures 5A-B).
Regarding instant claim 16, Latham, in view of McCarthy, teaches the method of claim 15. Latham further teaches the method further comprising determining whether the subject has full methylation of an expanded FMR1 allele (Page 13, paragraph [0132]; Page 16, paragraph [0137]; Table 5: FM=full methylation; Figures 5A-B).
Claim 12 is rejected under 35 U.S.C. 103 as being unpatentable over Latham (US 20120107824 A1) and McCarthy (MethylMeter(®): bisulfite-free quantitative and sensitive DNA methylation profiling and mutation detection in FFPE samples, Epigenomics, June 2016, 8, 747-765; cited on the IDS filed November 21st, 2024), as evidenced by Applied Biosciences (DNA Fragment Analysis by Capillary Electrophoresis [online]. Applied Biosciences, [2014] [retrieved on February 13th, 2026]. Retrieved from: https://mugenomicscore.missouri.edu/PDF/fragment-analysis-chemistry-guide.pdf), as applied to claims 1-11 and 15-16 above, and further in view Panteleeva (Optimizing DNA Shearing for Next Generation Sequencing Library Preparation with the Bioruptor® [online]. diagenode, [2010] [retrieved on February 12th, 2026]. Retrieved from: https://www.diagenode.com/files/posters/Poster-BR-A3-V2_22_06_10.pdf).
Regarding instant claim 12, Latham, in view of McCarthy, teaches the method of claim 1. Both Latham (Page 11, paragraph [0105]) and McCarthy (Page 756, column 1, paragraph 5 to column 2, paragraph 1) further teach wherein the isolated total nucleic acid is fragmented.
Neither reference teaches wherein the isolated total nucleic acid is fragmented to fragments having a size ranging from 2.4-3.2 kilobase pairs (kb) prior to contacting the isolated total nucleic acid with the methyl binding protein.
Panteleeva, in a reasonably pertinent field, teaches on sonication optimization for specific fragment sizes, including mean fragment sizes of 3000bp (e.g. 3.0kb) (Abstract).
It would have been obvious to one of ordinary skill in the art, before the effective filing date of the claimed invention, to have modified the fragment of Latham, in view of McCarthy, with the sonication of specific fragment sizes of Panteleeva. Since Panteleeva teaches optimizing DNA shearing, which is reasonably pertinent to the fragmenting of Latham, in view of McCarthy, one of ordinary skill in the art would combine the two teachings with a reasonable expectation of success. One of ordinary skill in the art would have been motivated to make this modification sonication is ideal to fractionate chromatic for epigenetics studies and is capable of highly reproducible fragment distributions (Abstract).
Claim 13 is rejected under 35 U.S.C. 103 as being unpatentable over Latham (US 20120107824 A1) and McCarthy (MethylMeter(®): bisulfite-free quantitative and sensitive DNA methylation profiling and mutation detection in FFPE samples, Epigenomics, June 2016, 8, 747-765; cited on the IDS filed November 21st, 2024), as evidenced by Applied Biosciences (DNA Fragment Analysis by Capillary Electrophoresis [online]. Applied Biosciences, [2014] [retrieved on February 13th, 2026]. Retrieved from: https://mugenomicscore.missouri.edu/PDF/fragment-analysis-chemistry-guide.pdf), as applied to claims 1-11 and 15-16 above, and further in view of Rehli (US 20210371901 A1).
Regarding instant claim 13, Latham, in view of McCarthy, teaches the method of claim 1.
Neither reference teaches wherein the methyl binding protein is a bifunctional polypeptide comprising: (i) an Fc portion of an antibody; (ii) a short flexible peptide linker; and (iii) a DNA-binding domain of an MBD2 protein.
Rehli, in a reasonably pertinent field, teaches a methyl binding protein of a bifunctional polypeptide comprising: (i) an Fc portion of an antibody (Page 18, paragraph [0150]); (ii) a short flexible peptide linker (Page 18, paragraph [0150]); and (iii) a DNA-binding domain of an MBD2 protein (Page 18, paragraph [0150]).
It would have been obvious to one of ordinary skill in the art, before the effective filing date of the claimed invention, to have modified the methyl binding protein of Latham, in view of McCarthy, with the methyl binding protein of Rehli. Since Rehli teaches on methods for detecting methylated DNA, which is reasonably pertinent to the method of Latham, in view of McCarthy, one of ordinary skill in the art would combine the two teachings with a reasonable expectation of success. One of ordinary skill in the art would have been motivated to make this modification because a bifunctional polypeptide as described has a high affinity and high avidity to methylated DNA (Rehli, Page 5, paragraph [0037]).
Claim 18 is rejected under 35 U.S.C. 103 as being unpatentable over McCarthy (MethylMeter(®): bisulfite-free quantitative and sensitive DNA methylation profiling and mutation detection in FFPE samples, Epigenomics, June 2016, 8, 747-765; cited on the IDS filed November 21st, 2024), in view of Panteleeva (Optimizing DNA Shearing for Next Generation Sequencing Library Preparation with the Bioruptor® [online]. diagenode, [2010] [retrieved on February 12th, 2026]. Retrieved from: https://www.diagenode.com/files/posters/Poster-BR-A3-V2_22_06_10.pdf).
It is noted by the Examiner that everything claimed in the following system recites an intended use of each component and is merely generically claimed. As long as the components of the system have structure capable of performing the generic action of the component as claimed, any further description of the component is considered an “intended use” and does not impart any further structural limitation on the claimed subject matter.
Regarding instant claim 18, McCarthy teaches a system for determining methylation of an FMR1 gene in a sample from a subject comprising: a component for isolating total nucleic acid from the sample (Page 751, column 1, paragraph 5: centrifuge to spin down samples); a component for contacting a first portion of the fragmented nucleic acid with a methyl binding protein (Page 751, column 2, paragraph 5: “[m]agnetic beads bearing the MethylMagnet GST-MBD2 fusion protein were distributed to 1.7 ml microcentrifuge tubes”); a component for collecting a fraction of the nucleic acid that is enriched for methylated nucleic acid and a fraction of the nucleic acid that is enriched for unmethylated nucleic acid (Page 751, column 2, paragraph 5 to Page 752, column 1, paragraph 1: “beads were pelleted with a magnet”); a component to perform PCR amplification of the FMR1 gene separately for the nucleic acid that is enriched for methylated nucleic acid and the fraction of the nucleic acid that is enriched for unmethylated nucleic acid (Page 754, column 1, paragraph 2: qPCR machine); and a component to determine if FMR1 gene sequences are present in the nucleic acid fraction enriched for methylated nucleic acid and/or the nucleic acid fraction enriched for unmethylated nucleic acid or both (Page 754, column 1, paragraph 2: FAM-BHQ1-labeled molecular beacon).
McCarthy does not teach a component for fragmenting the isolated total nucleic acid to a specific size range.
Panteleeva, in a reasonably pertinent field, teaches on a sonicator and sonication optimization for specific fragment sizes, including mean fragment sizes of 3000bp (e.g. 3.0kb) (Abstract: “Bioruptor sonicator”).
It would have been obvious to one of ordinary skill in the art, before the effective filing date of the claimed invention, to have modified the system of McCarthy with the sonicator for fragmenting nucleic acid to a specific size range of Panteleeva. Since Panteleeva teaches optimizing DNA shearing, which is reasonably pertinent to the system of McCarthy, one of ordinary skill in the art would combine the two teachings with a reasonable expectation of success. One of ordinary skill in the art would have been motivated to make this modification because sonicators are ideal to fractionate chromatic for epigenetics studies and is capable of highly reproducible fragment distributions (Abstract).
Conclusion
All claims stand rejected.
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/ALLISON E SCHLOOP/Examiner, Art Unit 1683
/Robert T. Crow/Primary Examiner, Art Unit 1683