CTNF 18/251,604 CTNF 78701 DETAILED ACTION Status of the Application Claims 1-20 are pending. 07-03-aia AIA 15-10-aia The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA. Applicant’s election without traverse of Group 6, claims 1, 4, 6-10, drawn in part to an in vitro method for enzymatic preparation of hexanoyl-CoA, and the combination of the enzymes acetyl-CoA acetyltransferase (ThlA), β-hydroxybutyryl-CoA dehydrogenase(Hbd1), 3-hydroxybutyryl-CoA dehydratase (Crt), trans-enoyl-CoA reductase (Ter), and thioesterase 1 (TesA), as submitted in a communication filed on 4/27/2026 is acknowledged. 08-06 AIA Claim s 2-3, 5, 11-20 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention , there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 4/27/2026 . Claims 1, 4, 6-10 are at issue and will be examined to the extent they encompass the elected invention . Specification The title of the invention is not descriptive. A new title is required that is clearly indicative of the invention to which the claims are directed. The claims under consideration are directed to a method. Appropriate correction is required. 07-29-04 AIA The disclosure is objected to because it contains an embedded hyperlink and/or other form of browser-executable code. See page 45, Table 1, two instances. Applicant is required to delete the embedded hyperlink and/or other form of browser-executable code. See MPEP § 608.01. Priority Acknowledgment is made of a claim for domestic priority under 35 U.S.C. 119(e) to provisional application No. 63/109,686 filed on 11/04/2020. This is the US national application which entered the national stage from PCT/US21/72188 filed on 11/02/2021. Information Disclosure Statement The information disclosure statements (IDS) submitted on 7/13/2023 and 3/20/2025 are acknowledged. The submissions are in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statements are being considered by the examiner. Drawings The drawings submitted on 5/3/2023 have been reviewed and are accepted by the Examiner for examination purposes. Claim Rejections - 35 USC § 112(b) or Second Paragraph (pre-AIA) 07-30-02 AIA 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. 07-34-01 AIA Claim s 1, 4, 6-10 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 pre-AIA the applicant regards as the invention. Claim 1 (claims 4, 6-10 dependent thereon) is indefinite in the recitation of “…the method comprising: (a) reacting…(b) expressing the translation…..one or more enzymes, (c) combining the cell-free protein….” for the following reasons. Since there is no “or” or “and” between steps (b) and (c), it is unclear if the steps (a)-(c) are in the alternative (i.e., “or”) or if the method requires all the steps (i.e., “and”). For examination purposes, it will be assumed that the method requires steps (a), (b) and (c). Correction is required. Claim 6 is indefinite in the recitation of “host strain…comprises Escherichia coli “ for the following reasons. While a host strain can be E. coli , it is unclear as to how a strain (unicellular microorganism) can comprise an E. coli cell. For examination purposes, it will be assumed that the claim recites “host strain… is Escherichia coli “. Correction is required. Claim 7 indefinite in the recitation of “host strain comprises one or more of E. coli strain BL21, JS07…and JC01” for the following reasons. It is unclear as to how a strain (unicellular microorganism) can comprise one or more E. coli strains. For examination purposes, it will be assumed that the claim recites “host strain is E. coli strain BL21, JS07… or JC01”. Correction is required. Claim 8 is indefinite in the recitation of “wherein the host strain comprises JS07”. While a host strain can b e E. coli JS07, it is unclear as to how a strain (unicellular microorganism) can comprise an E. coli JS07 cell. For examination purposes, it will be assumed that the claim recites “host strain… is Escherichia coli JS07“. Correction is required. Claim 9 is indefinite in the recitation of “wherein the cell-free protein synthesis reaction and the secondary reaction are in separate reaction vessels” for the following reasons. According to claim 1, the secondary reaction mixture is the result of combining the cell free protein synthesis reaction mixture with feedstock. Therefore, the secondary reaction mixture would have to be in the same vessel as the cell free protein synthesis reaction mixture. For examination purposes, it will be assumed that claim 9 is a duplicate of claim 1. Correction is required. Claim 10 is indefinite in the recitation of “wherein the cell-free protein synthesis reaction and the secondary reaction are in the same reaction vessel” for the following reasons. There is no antecedent basis for the cell-free protein synthesis reaction or the secondary reaction. For examination purposes, it will be assumed that claim 10 is a duplicate of claim 1. Correction is required. When amending the claims, applicant is advised to carefully review all examined claims and make the necessary changes to ensure proper antecedent basis and dependency. Claim Rejections - 35 USC § 112(a) or First Paragraph (pre-AIA) 07-30-01 AIA 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. 07-31-01 AIA Claim s 1, 4, 6-10 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 pre-AIA the inventor(s), at the time the application was filed, had possession of the claimed invention. As stated in MPEP 2111.01, during examination, the claims must be interpreted as broadly as their terms reasonably allow. Claims 1, 4, 6-10 are directed in part to an in vitro method for producing hexanoic acid, wherein said method comprises reacting a cell-free protein synthesis reaction mixture with cell-free protein synthesis reagents, wherein said cell-free protein synthesis reaction mixture comprises a cellular extract from a genus of host cells and translation templates encoding a genus of acetyl-CoA acetyltransferases having any structure, a genus of β-hydroxybutyryl-CoA dehydrogenases having any structure, a genus of 3-hydroxybutyryl-CoA dehydratases having any structure, a genus of trans-enoyl-CoA reductases having any structure, and/or a genus of thioesterases having any structure. See Claim Rejections - 35 USC § 112(b) or Second Paragraph (pre-AIA) for claim interpretation. In University of California v. Eli Lilly & Co. , 43 USPQ2d 1938, the Court of Appeals for the Federal Circuit has held that “A written description of an invention involving a chemical genus, like a description of a chemical species, ‘requires a precise definition, such as by structure, formula, [or] chemical name,’ of the claimed subject matter sufficient to distinguish it from other materials”. As indicated in MPEP § 2163, the written description requirement for a claimed genus may be satisfied through sufficient description of a representative number of species by actual reduction to practice, reduction to drawings, or by disclosure of relevant, identifying characteristics, i.e., structure or other physical and/or chemical properties, by functional characteristics coupled with a known or disclosed correlation between function and structure, or by a combination of such identifying characteristics, sufficient to show that Applicant was in possession of the claimed genus. In addition, MPEP § 2163 states that a representative number of species means that the species which are adequately described are representative of the entire genus. Thus, when there is substantial variation within the genus, one must describe a sufficient variety of species to reflect the variation within the genus. There is no structural limitation with respect to the members of the genus of enzymes required by the claimed method. While the specification in the instant application discloses the structure of a limited number of species of the recited genus of acetyl-CoA acetyltransferases, β-hydroxybutyryl-CoA dehydrogenases, 3-hydroxybutyryl-CoA dehydratases, trans-enoyl-CoA reductases, and thioesterases, it provides no clue as to the structural elements required in any acetyl-CoA acetyltransferase, β-hydroxybutyryl-CoA dehydrogenase, 3-hydroxybutyryl-CoA dehydratase, trans-enoyl-CoA reductase, and thioesterase that can be used to make hexanoic acid. In addition, while some of the claims require the cellular extract of a genus of host cells from any species, the specification and the prior art are silent as to whether any host cell’s extract in combination with the recited enzymes could be used to produce hexanoic acid. No disclosure of a structure/function correlation has been provided which would allow one of skill in the art to recognize which proteins have the recited enzymatic activities. The claims encompass a large genus of proteins which are structurally unrelated. A sufficient written description of a genus of polypeptides may be achieved by a recitation of a representative number of polypeptides defined by their amino acid sequence or a recitation of structural features common to members of the genus, which features constitute a substantial portion of the genus. However, in the instant case, there is no recited structural feature which is representative of all the members of the genus of enzymes recited in the claims. Furthermore, while one could argue that the few species disclosed are representative of the structure of all the members of the genus of enzymes required, it is noted that the art teaches several examples of how even highly structurally homologous polypeptides can have different enzymatic activities. For example, Witkowski et al. (Biochemistry 38:11643-11650, 1999) teach that one conservative amino acid substitution transforms a β-ketoacyl synthase into a malonyl decarboxylase and completely eliminates β-ketoacyl synthase activity. Tang et al. (Phil Trans R Soc B 368:20120318, 1-10, 2013) teach that two Dehalobacter reductive dehalogenases, CfrA and DcrA, having 95.2% sequence identity to teach other have exclusively different substrate (Abstract; page 7, left column, Discussion, CfrA and DcrA). Seffernick et al. (J. Bacteriol. 183(8):2405-2410, 2001) teach that two naturally occurring Pseudomonas enzymes having 98% amino acid sequence identity catalyze two different reactions: deamination and dehalogenation, therefore having different function. Therefore, since minor structural differences may result in changes affecting function, and no additional information correlating structure with the desired functional characteristics has been provided, one cannot reasonably conclude that the few species disclosed are representative of the structure of all the enzymes required by the claimed method. Due to the fact that the specification only discloses a limited number of species of the genus of acetyl-CoA acetyltransferases, β-hydroxybutyryl-CoA dehydrogenases, 3-hydroxybutyryl-CoA dehydratases, trans-enoyl-CoA reductases, and thioesterases required by the claimed method, E. coli as the only host strain to provide a cellular extract that could sustain hexanoic acid synthesis in vitro , and the lack of description of any additional species by any relevant, identifying characteristics or properties, one of skill in the art would not recognize from the disclosure that Applicant was in possession of the claimed invention . 07-31-02 AIA Claim s 7-8 rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA), first paragraph, as failing to comply with the enablement requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to enable one skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention. The invention appears to employ novel strains (i.e., E. coli JS07, MB263, MB263sucD, and JC01). Since the strains are essential to the claimed invention, they must be obtainable by a repeatable method set forth in the specification or otherwise be readily available to the public. There is no indication that E. coli JS07, MB263, MB263sucD, and JC01 are readily available to the public. The enablement requirements of 35 U.S.C. §112 may be satisfied by a deposit of the strains. The specification does not disclose a repeatable process to obtain the strains. Accordingly, it is deemed that a deposit of these strains should have been made in accordance with 37 CFR 1.801-1.809. If a deposit was made under the terms of the Budapest Treaty, then an affidavit or declaration by applicants, or a statement by an attorney of record over his or her signature and registration number, stating that the specific strains have been deposited under the Budapest Treaty and that the strains will be available to the public under the conditions specified in 37 CFR 1.808, would satisfy the deposit requirement made herein. If a deposit has not been made under the Budapest treaty, then in order to certify that the deposit meets the criteria set forth in 37 CFR 1.801-1.809, Applicant may provide assurance or compliance by an affidavit or declaration, or by a statement by an attorney of record over his or her signature and registration number, showing that: a. during the pendency of this application, access to the invention will be afforded to the Commissioner upon request; b. upon granting of the patent the strains will be available to the public under the conditions specified in 37 CFR 1.808; c. the deposit will be maintained in a public repository for a period of 30 years or 5 years after the last request or for the effective life of the patent, whichever is longer; and d. the deposit will be replaced if it should ever become non-viable . 07-31-03 Claim s 1, 4, 6-10 are rejected under 35 U.S.C. 112, first paragraph, because the specification, while being enabling for an in vitro method for producing hexanoic acid, wherein said method comprises (a) reacting a cell-free protein synthesis reaction mixture with cell-free protein synthesis reagents, wherein said cell-free protein synthesis reaction mixture comprises a cellular extract from E. coli , a translation template for the C. kluyveri th1A gene, which encodes an acetyl-CoA acetyltransferase, a translation template for the C. kluyveri hbd1 gene, which encodes a β-hydroxybutyryl-CoA dehydrogenase, a translation template for the C. acetobutylicum crt gene, which encodes a 3-hydroxybutyryl-CoA dehydratase, a translation template for the T . denticola ter gene, which encodes a trans-enoyl-CoA reductase, and a translation template for the E. coli tesA gene, which encodes a thioesterase, (b) expressing the translation templates in the cell-free protein synthesis reaction mixture to obtain the C. kluyveri acetyl-CoA acetyltransferase, the C. kluyveri β-hydroxybutyryl-CoA dehydrogenase, the C. acetobutylicum 3-hydroxybutyryl-CoA dehydratase, the T . denticola trans-enoyl-CoA reductase, and the E. coli thioesterase 1, and (c) combining the cell-free protein synthesis reaction mixture comprising the C. kluyveri acetyl-CoA acetyltransferase, the C. kluyveri β-hydroxybutyryl-CoA dehydrogenase, the C. acetobutylicum 3-hydroxybutyryl-CoA dehydratase, the T . denticola trans-enoyl-CoA reductase, and the E. coli thioesterase 1 with feedstock that comprises glucose to form a secondary reaction mixture, wherein the feedstock reacts in the presence of the C. kluyveri acetyl-CoA acetyltransferase, the C. kluyveri β-hydroxybutyryl-CoA dehydrogenase, the C. acetobutylicum 3-hydroxybutyryl-CoA dehydratase, the T . denticola trans-enoyl-CoA reductase, and the E. coli thioesterase 1 to produce hexanoic acid as well as the intermediates acetyl-CoA, acetoacetyl-CoA, 3-hydroxybutyryl-CoA, hex-(2E)-enoyl-CoA and hexanoyl-CoA , does not reasonably provide enablement for an in vitro method for producing hexanoic acid, wherein said method comprises reacting a cell-free protein synthesis reaction mixture with cell-free protein synthesis reagents, wherein said cell-free protein synthesis reaction mixture comprises a cellular extract from any host cell and translation templates encoding any acetyl-CoA acetyltransferase, any β-hydroxybutyryl-CoA dehydrogenase, any 3-hydroxybutyryl-CoA dehydratase, any trans-enoyl-CoA reductase, and/or any thioesterase 1 . The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention commensurate in scope with these claims. Factors to be considered in determining whether undue experimentation is required are summarized in In re Wands (858 F.2d 731, 737, 8 USPQ2nd 1400 (Fed. Cir. 1988)) as follows: 1) quantity of experimentation necessary, 2) the amount of direction or guidance presented, 3) the presence and absence of working examples, 4) the nature of the invention, 5) the state of prior art, 6) the relative skill of those in the art, 7) the predictability or unpredictability of the art, and 8) the breadth of the claims. The factors which have led the Examiner to conclude that the specification fails to teach how to make and/or use the claimed invention without undue experimentation, are addressed in detail below. The breadth of the claims . Claims 1, 4, 6-10 broadly encompass an in vitro method for producing hexanoic acid, wherein said method comprises reacting a cell-free protein synthesis reaction mixture with cell-free protein synthesis reagents, wherein said cell-free protein synthesis reaction mixture comprises a cellular extract from any host cell and translation templates encoding any acetyl-CoA acetyltransferase, any β-hydroxybutyryl-CoA dehydrogenase, any 3-hydroxybutyryl-CoA dehydratase, any trans-enoyl-CoA reductase, and/or any thioesterase 1. The enablement provided is not commensurate in scope with the claims due to the extremely large number of proteins of unknown structure encompassed by the claims, the lack of information regarding the structural elements required in any of the enzymes recited, and the unknown host cells that could be used to obtain a cellular extract to make hexanoic acid. In the instant case, the specification enables an in vitro method for producing hexanoic acid, wherein said method comprises (a) reacting a cell-free protein synthesis reaction mixture with cell-free protein synthesis reagents, wherein said cell-free protein synthesis reaction mixture comprises a cellular extract from E. coli , a translation template for the C. kluyveri th1A gene, which encodes an acetyl-CoA acetyltransferase, a translation template for the C. kluyveri hbd1 gene, which encodes a β-hydroxybutyryl-CoA dehydrogenase, a translation template for the C. acetobutylicum crt gene, which encodes a 3-hydroxybutyryl-CoA dehydratase, a translation template for the T . denticola ter gene, which encodes a trans-enoyl-CoA reductase, and a translation template for the E. coli tesA gene, which encodes a thioesterase, (b) expressing the translation templates in the cell-free protein synthesis reaction mixture to obtain the C. kluyveri acetyl-CoA acetyltransferase, the C. kluyveri β-hydroxybutyryl-CoA dehydrogenase, the C. acetobutylicum 3-hydroxybutyryl-CoA dehydratase, the T . denticola trans-enoyl-CoA reductase, and the E. coli thioesterase 1, and (c) combining the cell-free protein synthesis reaction mixture comprising the C. kluyveri acetyl-CoA acetyltransferase, the C. kluyveri β-hydroxybutyryl-CoA dehydrogenase, the C. acetobutylicum 3-hydroxybutyryl-CoA dehydratase, the T . denticola trans-enoyl-CoA reductase, and the E. coli thioesterase 1 with feedstock that comprises glucose to form a secondary reaction mixture, wherein the feedstock reacts in the presence of the C. kluyveri acetyl-CoA acetyltransferase, the C. kluyveri β-hydroxybutyryl-CoA dehydrogenase, the C. acetobutylicum 3-hydroxybutyryl-CoA dehydratase, the T . denticola trans-enoyl-CoA reductase, and the E. coli thioesterase 1 to produce hexanoic acid as well as the intermediates acetyl-CoA, acetoacetyl-CoA, 3-hydroxybutyryl-CoA, hex-(2E)-enoyl-CoA and hexanoyl-CoA. The amount of direction or guidance presented and the existence of working examples . The specification discloses an in vitro method to produce hexanoic acid which uses an E. coli cell extract in combination with an acetyl-CoA acetyltransferase encoded by the C. kluyveri th1A gene, the β-hydroxybutyryl-CoA dehydrogenase encoded by the C. kluyveri hbd1 gene, the 3-hydroxybutyryl-CoA dehydratase encoded by the C. acetobutylicum crt gene, the trans-enoyl-CoA reductase encoded by the T . denticola ter gene, and the thioesterase encoded by the E. coli tesA gene as a working example. However, the specification fails to provide any clue as to the structural elements required in any acetyl-CoA acetyltransferase, β-hydroxybutyryl-CoA dehydrogenase, 3-hydroxybutyryl-CoA dehydratase, trans-enoyl-CoA reductase, and thioesterase. No correlation between structure and function has been presented. The state of prior art, the relative skill of those in the art, and the predictability or unpredictability of the art . The amino acid sequence of a polypeptide determines its structural and functional properties. While the art discloses a limited number of acetyl-CoA acetyltransferases, β-hydroxybutyryl-CoA dehydrogenases, 3-hydroxybutyryl-CoA dehydratases, trans-enoyl-CoA reductases, and thioesterases, neither the specification nor the art provide a correlation between structure and function such that one of skill in the art can envision the structure of any of the enzymes required by the claimed method. The art clearly teaches that (a) determining function based solely on structural homology, and (b) modification of a protein’s amino acid sequence to obtain the desired activity without any guidance/knowledge as to which amino acids in a protein are tolerant of modification and which ones are conserved are highly unpredictable. For example, Singh et al. (Current Protein and Peptide Science 19(1):5-15, 2018) disclose different protein engineering approaches and state that despite the availability of an ever-growing database of protein structures and highly sophisticated computational algorithms, protein engineering is still limited by the incomplete understanding of protein functions, folding, flexibility and conformational changes (page 11, left column, last paragraph). Sadowski et al. (Current Opinion in Structural Biology 19:357-362, 2009) teach that much of the problem in assigning function from structure comes from functional convergence, where although a stable structure is required to perform many functions it is not always necessary to adopt a particular structure to carry out a particular function (page 357, right column, first full paragraph). Sadowski et al. further explain that the unexpected and significant difficulties of predicting function from structure show that the potential of structural models for providing novel functional annotations has not yet fully realized. Sadowski et al. also states that while a few successes have been achieved which required manual intervention, the ability to vary the requirements for specificity in prediction means that it is difficult to determine how useful the end result may be for the user (page 361, left column, first full paragraph). The teachings of Singh et al. and Sadowski et al. are further supported by the teachings of Witkowski et al., Tang et al. and Seffernick et al. already discussed above, where it is shown that even small amino acid changes result in enzymatic activity changes. The quantity of experimentation required to practice the claimed invention based on the teachings of the specification. While methods of generating or isolating variants of a polypeptide and enzymatic assays were known in the art at the time of the invention, it was not routine in the art to screen by a trial and error process for an essentially infinite number of proteins to find a protein with acetyl-CoA acetyltransferase activity, β-hydroxybutyryl-CoA dehydrogenase activity, 3-hydroxybutyryl-CoA dehydratase activity, trans-enoyl-CoA reductase activity, and thioesterase activity. Similarly, it was not routine in the art to screen by a trial and error process for an essentially infinite number of combinations of host cell extracts and enzymes to determine which ones can be used to produce hexanoic acid. In the absence of (i) a rational and predictable scheme for selecting those proteins most likely to have the desired functional features, and/or (ii) a correlation between structure and the desired enzymatic activity, one of skill in the art would have to test an essentially infinite number of proteins to determine which ones have the desired functional characteristics. Therefore, taking into consideration the extremely broad scope of the claim, the lack of guidance, the amount of information provided, the lack of knowledge about a correlation between structure and the desired function, and the high degree of unpredictability of the prior art in regard to structural changes and their effect on function, one of ordinary skill in the art would have to go through the burden of undue experimentation in order to practice the claimed invention. Thus, Applicant has not provided sufficient guidance to enable one of ordinary skill in the art to make and use the invention in a manner reasonably correlated with the scope of the claims. Claim Rejections - 35 USC § 102 (AIA) 07-07-aia AIA 07-07 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 – 07-08-aia AIA (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. 07-15 AIA Claim s 1, 4, 6-7, 9-10 are rejected under 35 U.S.C. 102( a)(1 ) as being anticipated by Karim et al. (Nature Chemical Biology 16:912-919, published on-line 6/15/2020; cited in the IDS) . Claims 1, 4, 6-7, 9-10 are directed in part to an in vitro method for producing acetoacetyl-CoA and 3-hydroxybutyryl-CoA, wherein said method comprises (a) reacting a cell-free protein synthesis reaction mixture with cell-free protein synthesis reagents, wherein said cell-free protein synthesis reaction mixture comprises a cellular extract from E. coli BL21, a translation template encoding an acetyl-CoA acetyltransferase, and a translation template encoding a β-hydroxybutyryl-CoA dehydrogenase, (b) expressing the translation templates in the cell-free protein synthesis reaction mixture to obtain the acetyl-CoA acetyltransferase and the β-hydroxybutyryl-CoA dehydrogenase, and (c) combining the cell-free protein synthesis reaction mixture comprising the acetyl-CoA acetyltransferase and the β-hydroxybutyryl-CoA dehydrogenase, with feedstock that comprises glucose to form a secondary reaction mixture, wherein the feedstock reacts in the presence of the acetyl-CoA acetyltransferase and the β-hydroxybutyryl-CoA dehydrogenase, to produce acetoacetyl-CoA and 3-hydroxybutyryl-CoA, wherein the method can comprise the addition of NAD+ and coenzyme A to the secondary reaction mixture. See Claim Rejections - 35 USC § 112(b) or Second Paragraph (pre-AIA) for claim interpretation. Karim et al. teach an in vitro method for 3-HB (3-hydroxybutyrate) biosynthesis which uses a cell-free system (Abstract). The method of Karim et al. produces the intermediates acetoacetyl-CoA and 3-hydroxybutyryl-CoA (Figure 2a). Karim et al. teach the use of an E. coli Bl21 cell lysate (Methods-Bacterial strains and plasmids), wherein said E. coli Bl21 expresses a C. acetobutylicum acetyl-CoA acetyltransferase (CacThl) and a C. kluyveri β-hydroxybutyryl-CoA dehydrogenase (ClkHbd1; page 913, right column, first full paragraph). Thus, the E. coli Bl21 cell lysate comprises translation templates that encode the C. acetobutylicum acetyl-CoA acetyltransferase (CacThl) and the C. kluyveri β-hydroxybutyryl-CoA dehydrogenase. Karim et al. teach reacting a solution comprising the E. coli cell lysate with cell-free protein reagents to express the translation templates and obtain the enzymes. Karim et al. teach further adding a solution comprising glucose as well as NAD and CoA to carry out the reactions catalyzed by the enzymes and produce the intermediates (Methods-iPROBE reactions). Therefore, the method of Karim et al. anticipate the instant claims as written/interpreted . 07-06 AIA 15-10-15 In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status. Conclusion No claim is in condition for allowance. 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. Applicant is advised that any Internet email communication by the Examiner has to be authorized by Applicant in written form . See MPEP § 502.03 (II). Without a written authorization by Applicant in place, the USPTO will not respond via Internet email to any Internet correspondence which contains information subject to the confidentiality requirement as set forth in 35 U.S.C. 122. Sample written authorization language can be found in MPEP § 502.03 (II). An Authorization for Internet Communications in a Patent Application or Request to Withdraw Authorization for Internet Communications form (SB/439) can be found at https://www.uspto.gov/patent/forms/ forms-patent- applications-filed-or-after-september-16-2012, which can be electronically filed. Any inquiry concerning this communication or earlier communications from the examiner should be directed to DELIA M RAMIREZ, Ph.D., whose telephone number is (571) 272-0938. The examiner can normally be reached on Monday-Friday from 8:30 AM to 5:00 PM. If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Robert B. Mondesi, can be reached at (408) 918-7584. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300. /DELIA M RAMIREZ/Primary Examiner, Art Unit 1652 DR May 28, 2026 Application/Control Number: 18/251,604 Page 2 Art Unit: 1652 Application/Control Number: 18/251,604 Page 3 Art Unit: 1652 Application/Control Number: 18/251,604 Page 4 Art Unit: 1652 Application/Control Number: 18/251,604 Page 5 Art Unit: 1652 Application/Control Number: 18/251,604 Page 6 Art Unit: 1652 Application/Control Number: 18/251,604 Page 7 Art Unit: 1652 Application/Control Number: 18/251,604 Page 9 Art Unit: 1652 Application/Control Number: 18/251,604 Page 10 Art Unit: 1652 Application/Control Number: 18/251,604 Page 11 Art Unit: 1652 Application/Control Number: 18/251,604 Page 12 Art Unit: 1652 Application/Control Number: 18/251,604 Page 13 Art Unit: 1652 Application/Control Number: 18/251,604 Page 14 Art Unit: 1652 Application/Control Number: 18/251,604 Page 15 Art Unit: 1652 Application/Control Number: 18/251,604 Page 16 Art Unit: 1652
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