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 an oligonucleotide as the inhibitor in claims 21 and 29, specifically an shRNA as the oligonucleotide in claims 22 and 30, more specifically SEQ ID NO: 1 as the shRNA in claims 23,24,31 and 32 in the reply filed on 11/17/2025 is acknowledged. Upon further consideration, in light of discovered prior art, the species election of shRNA as the inhibitor of EXT1 has been expanded to include guide RNA . Regarding the elected sequence, SEQ ID NO: 1 is free of prior art and therefore the species election has been expanded to t he oligonucleotide o f SEQ ID NO: 2. Claims 16-34 are under examination. Priority This application is a 371 of PCT/EP2021/054190, filed 02/19/2021, and claims foreign priority to EP20158875.3, filed 02/21/2020. Claim Objections Claim s 16- 34 are objected to because of the following informalities: Independent claim 16 recites “EXT1” in step a) and independent claim 26 recites “EXT1” in step c) . For purposes of clarity, at least the first recitation of the name of a gene should be fully written out rather than the acronym , “EXT1” . Claims 17-25 are objected to because they depend from claim 16 and either do not correct the issue or also recite “EXT1”, and claims 27-34 depend from claim 26 and either do not correct the issue or recite “EXT1”. Appropriate correction is required. Claims 23,24,31 and 32 are objected to because of the following informalities: The SEQ ID NOs: recited in claims 23,24,31 and 32 are bolded. Appropriate correction is required. Claim 26 is objected to because of the following informalities: step b) in claim 26 has a period after “entity” rather than a semicolon. Appropriate correction is required. Specification The disclosure is objected to because it contains an embedded hyperlink and/or other form of browser-executable code. Applicant is required to delete the embedded hyperlink and/or other form of browser-executable code; references to websites should be limited to the top-level domain name without any prefix such as http:// or other browser-executable code. See MPEP § 608.01. See at least pages 31,43 and 54-56 which recite https or www. Improper Markush Grouping Rejection Claim s 21 and 29 are rejected on the basis that it contains an improper Markush grouping of alternatives. See In re Harnisch , 631 F.2d 716, 721-22 (CCPA 1980) and Ex parte Hozumi , 3 USPQ2d 1059, 1060 (Bd. Pat. App. & Int. 1984). A Markush grouping is proper if the alternatives defined by the Markush group (i.e., alternatives from which a selection is to be made in the context of a combination or process, or alternative chemical compounds as a whole) share a “single structural similarity” and a common use. A Markush grouping meets these requirements in two situations. First, a Markush grouping is proper if the alternatives are all members of the same recognized physical or chemical class or the same art-recognized class, and are disclosed in the specification or known in the art to be functionally equivalent and have a common use. Second, where a Markush grouping describes alternative chemical compounds, whether by words or chemical formulas, and the alternatives do not belong to a recognized class as set forth above, the members of the Markush grouping may be considered to share a “single structural similarity” and common use where the alternatives share both a substantial structural feature and a common use that flows from the substantial structural feature. See MPEP § 2117. The Markush grouping of “wherein said inhibitor of the EXT1 expression and/or activity is selected from a group comprising an oligonucleotide, an aptamer, an oligopeptide, a polypeptide, a chemical compound and an analog thereof” is improper because the alternatives defined by the Markush grouping do not share both a single structural similarity and a common use for the following reasons: all of the members of the Markush grouping above belong to different chemical or art-recognized classes and do not share a single structural similarity. Peptides and polypeptides have different structures than oligonucleotides and aptamers, and “chemical compounds and an analog thereof” also do not share the same structure as oligonucleotides, aptamers and peptides/polypeptides. In addition, “chemical compounds and an analog thereof” encompasses any chemical compound or analog thereof which may not have a single structural similarity of all of the possible chemical compounds encompassed by the genus. To overcome this rejection, Applicant may set forth each alternative (or grouping of patentably indistinct alternatives) within an improper Markush grouping in a series of independent or dependent claims and/or present convincing arguments that the group members recited in the alternative within a single claim in fact share a single structural similarity as well as a common use. 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. Clai ms 16-25 and 28-30 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. Regarding claim 16 , the phrase "preferably" in step b) renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d). Claims 17-25 are included in the rejection because they depend from claim 16 and do not correct the issue. Regarding claim s 18 ,21-24 and 28-30, the claims recite the following phrases including “selected in a group comprising”, “selected from a group comprising”, “selected from a group comprising or consisting of”. Regarding Markush language and grouping, MPEP 2173.05(h) states: A Markush grouping is a closed group of alternatives, i.e., the selection is made from a group "consisting of" (rather than "comprising" or "including") the alternative members. Abbott Labs., 334 F.3d at 1280, 67 USPQ2d at 1196. If a Markush grouping requires a material selected from an open list of alternatives (e.g., selected from the group "comprising" or "consisting essentially of" the recited alternatives), the claim should generally be rejected under 35 U.S.C. 112(b) as indefinite because it is unclear what other alternatives are intended to be encompassed by the claim. See In re Kiely, 2022 USPQ2d 532 at 2* (Fed. Cir. 2022) (each independent claim recites "a selection from the group comprising a person, an animal, an animated character, a creature, an alien, a toy, a structure, a vegetable, and a fruit." … (emphasis added). "Given the breadth of variation among the specified alternatives and the use of the open-ended word ’comprising’ to define the scope of the list, we affirm the Board's conclusion that the pending claims recite improper Markush language and are indefinite under § 112(b) . "). If a claim is intended to encompass combinations or mixtures of the alternatives set forth in the Markush grouping, the claim may include qualifying language preceding the recited alternatives (such as "at least one member" selected from the group), or within the list of alternatives (such as "or mixtures thereof"). Id. at 1281. See also MPEP § 2111.03 . Therefore, claims 18,21-24 and 28-30 do not recite a closed group of alternatives as required by a Markush grouping as “selected in a group comprising ”, “selected from a group comprising ” and “selected from a group comprising or consisting of” is open language. Written Description Rejection 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. Claim s 16-34 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. C laim s 16-18 and 25 are directed to encompass a method for production of a biological entity (polypeptide or viral particle) in a cell comprising a step of providing a cell population having at least depleted EXT1 expression and/or activity by any means, and claim 19 recites the cell comprises a partial or total knockout of the EXT1 gene. Claims 20,26-28,33 and 34 recite depleted EXT1 expression and/or activity is obtained by treatment of said cell with an inhibitor of EXT1 expression and/or activity, which encompasses any type of EXT1 inhibitor. Claims 21 and 29 recite the inhibitor of EXT1 expression or activity is an oligonucleotide, an aptamer, an oligopeptide, a polypeptide, a chemical compound and an analog thereof, and claims 22 and 30 recite the inhibitor of the EXT1 expression is antisense RNA, a miRNA, a guide RNA, a siRNA and a shRNA. Claims 23,24,31 and 32 recite specific sequences of the EXT1 inhibitor. Therefore, the instant claims recite a method of producing a biological entity (a polypeptide or viral particle) comprising a step of providing a cell population produced by a genus of means for depleting EXT1 expression and/or activity or by using a large genus of EXT1 inhibitors , which only correspond in some undefined way to specifically instantly disclosed shRNA and siRNAs. Regarding the state of the art, Tokyo (EP3604502, Published 5 Feb 2020), cited on an IDS , teaches methods for stably propagating virulent hand, foot and mouth disease viruses (paragraph 0007) and host cells therefore that preferably do not express the EXT1 gene (paragraphs 0011,0035), and preparing RD cells expressing no heparan sulfate by genome editing using CRISPR/Cas9 . Tokyo teaches the sequence of the guide RNA targeting the EXT1 gene was designed and shown below (paragraph 0072). Chen et al. (English translation of CN107058476, Published 18 August 2017), cited on an IDS, taught the construction of liver cancer cell lines that interfere with EXT1 expression using EXT1 shRNA sequences (page 8 of English translation). The EXT1 shRNA sequence 3-1 of Chen et al. shown below is 100% identical to the shRNA sequence of instant SEQ ID NO: 2. Chen et al. taught transfection of the EXT1 shRNA sequences into Hep3-B and Huh-7 cells, and that it successfully interfered with the expression of EXT1 in Hep3-B and Huh-7 liver cancer cell lines (page 9 and Fig. 4). Therefore, the state of the art teach virus production in cells by gene editing using a specific sequence of sgRNA targeting EXT1, and teaches shRNA sequences that interfere with EXT1 expression in cells. The instant specification generically contemplates inhibitors of EXT1 activity on pages 8-9,12-14. Pages 8-9 contemplate EXT1 inhibitors being an oligonucleotide, an aptamer, an oligopeptide, a polypeptide, a chemical compound and analog thereof, and the oligonucleotide is selected from an antisense RNA, a miRNA, a guide RNA, a siRNA a shRNA, and pages 12-14 contemplate oligopeptides of less than 50 amino acids, polypeptides of at least 50 amino acids, antibodies, antibody fragments, diabody, triabody , tetrabody , nanobody and analog thereof. The recitation “analog thereof” of a chemical compound encompasses many possible compounds, and is similar to the chemical compound that it is an analog of as possessing the same activity, but is structurally different. The specification has not provided any written description support for chemical compounds or any analogs of chemical compounds. The examples disclose EXT1 shRNAs of SEQ ID NOs: 1-24 on pages 31-33 and SEQ ID NOs: 33-53 on pages 56-57 and EXT1 siRNAs of SEQ ID NOs: 25-28 on page 34 . The results on page 55 for Example 2 show HEK293 shEXT1 produce approximately four times more lentiviral particles than the HEK293 shCTRL cells (Fig. 28) and approximately three times more AAV2 viral particles than HEK293 shCTRL cells (Fig. 29), as well as expressing 2.9 time more Notch1-Flag protein than control cells (Fig. 30) and 1.7 times more nano-luciferase enzyme than control cells (Fig. 31). However, it cannot be determined what the shRNA sequences were that had the function of depleting EXT1 expression and/or activity and production of the polypeptide or viral particle in the cell the cell population in order to determine the required structure that performs the recited functions. Example 4 on page 58 shows testing EXT1 targeting shRNA shown in Table 4, including #1 and #20 in HEK293 cells lines, which are then infected with lentiviral or AAV2 particles expressing Nano-luciferase enzyme or green fluorescent protein. Results of the western blot shows that not all shRNA sequences used are able to reduce the levels of EXT1 expression in HEK293 cells , and 8 out of 20 tested shRNAs were identified as able to induce reduction of EXT1 levels in cells, which are shRNA#3,4,7,11,12,16,18 and 20 which are SEQ ID NOs: 35,36,39,43,44, 48,50 and 52 respectively (Table 5, page 59). EXT1 knockdown confirmed cells were transduced with nanoluciferase expressing lentivirus or GRP expression AAV2 virus, and results were that shRNA #3 and #7 exhibited the highest productivity in both lentiviral and AAV systems (Page 59 and Fig. 33A-B) and other EXT1 knockdown cells lines showed significant productivity compared to control cells with shEXT1 #12,16 and 20 for lentiviruses (Fig. 33A) and shEXT1 #11,12,16 and 20 for AAV viruses (Fig. 33B). Therefore, the specification discloses that not all shRNA EXT1 sequences reduce EXT1 expression in cells and therefore the specification does not teach the required structure of EXT1 shRNA that carries out the recited functions, and also does not disclose the complete or partial structure of any other species of EXT1 inhibitors encompassed by the genus of EXT1 inhibitors. While claims 23,24,31 and 32 recite specific oligonucleotide sequences of the EXT1 inhibitor, the fact that the specification discloses that not all shRNA EXT1 sequences reduce EXT1 expression in cells adds to the lack of written description for what the necessary structure is to achieve the claimed functions, and that is why claims 23,24,31 and 32 are included in the rejection as only specific sequence as noted above were found to reduce EXT1 and result in the claimed functions. Therefore, the instant specification discloses how to produce a polypeptide or viral particle in a eukaryotic cell that has been depleted of EXT1 expression and/or activity using shRNAs described in Table 5, page 59 . Other than the EXT1 shRNAs in Table 5 on page 59 , n one of these EXT1 inhibitors meet the written description provision of 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA), first paragraph, due to lacking chemical structural information for what they are and chemical structures are highly variant and encompass a myriad of possibilities. The specification provides insufficient written description to support the genus encompassed by the claim. Note: MPEP 2163. Vas-Cath Inc. v. Mahurkar , 19 USPQ2d 1111, (Fed. Cir. 1991), makes clear that "applicant must convey with reasonable clarity to those skilled in the art that, as of the filing date sought, he or she was in possession of the invention. The invention is, for purposes of the 'written description' inquiry, whatever is now claimed. " (See page 1117.) The specification does not "clearly allow persons of ordinary skill in the art to recognize that [he or she] invented what is claimed." (See Vas-Cath at page 1116.) Univ. of Rochester v. G.D. Searle , 69 USPQ2d 1886, 1892 (CAFC 2004), further supports this by stating that: The appearance of mere indistinct words in a specification or a claim, even an original claim, does not necessarily satisfy that requirement. A description of an anti-inflammatory steroid, i.e., a steroid (a generic structural term) described even in terms of its functioning of lessening inflammation of tissues fails to distinguish any steroid from others having the same activity or function. A description of what a material does, rather than of what it is, usually does not suffice…. The disclosure must allow one skilled in the art to visualize or recognize the identity of the subject matter purportedly described. (Emphasis added). With the exception of the above specifically disclosed shRNAs in Table 5, page 59 , the specific EXT1 guide RNA of Tokyo, and the shRNA sequences of Chen et al., the skilled artisan cannot envision the detailed chemical structure of the encompassed EXT1 inhibitors with the recited function of depleting EXT1 expression and/or activity and resulting in production of a polypeptide or viral particle , regardless of the complexity or simplicity of the method of isolation. The structures of a polypeptide, oligonucleotide, aptamer and chemical compound are completely different and each encompasses thousands of possible species, and no structure-function correlation has been provided regarding what the required core structure is of an EXT1 inhibitor that has the recited function of depleting EXT1 expression and/or activity in a cell population and the function of producing a polypeptide or viral particle in a cell population. Adequate written description requires more than a mere statement that it is part of the invention and reference to a potential method for isolating it. The chemical structure itself is required. See Fiers v. Revel , 25 USPQ2d 1601, 1606 (Fed. Circ. 1993) and Amgen Inc. V. Chugai Pharmaceutical Co. Ltd., 18 USPQ2d 1016, (Fed. Cir. 1991). In Fiddes v. Baird , 30 USPQ2d 1481, 1483, (Bd. Pat. App. & Int. 1993), claims directed to mammalian FGF's were found unpatentable due to lack of written description for the broad class. The specification provided only the bovine sequence. Finally, University of California v. Eli Lilly and Co. , 43 USPQ2d 1398, 1404, 1405 (Fed. Cir. 1997) held that: ...To fulfill the written description requirement, a patent specification must describe an invention and do so in sufficient detail that one skilled in the art can clearly conclude that "the inventor invented the claimed invention." Lockwood v. American Airlines, Inc. , 107 F.3d 1565, 1572, 41 USPQ2d 1961, 1966 (Fed. Cir. 1997); In re Gosteli , 872 F.2d 1008, 1012, 10 USPQ2d 1614, 1618 (Fed. Cir. 1989) (" [T]he description must clearly allow persons of ordinary skill in the art to recognize that [the inventor] invented what is claimed."). Thus, an applicant complies with the written description requirement "by describing the invention, with all its claimed limitations, not that which makes it obvious," and by using "such descriptive means as words, structures, figures, diagrams, formulas, etc., that set forth the claimed invention." Lockwood , 107 F.3d at 1572, 41 USPQ2d at 1966. Furthermore, to the extent that a functional description can meet the requirement for an adequate written description, it can do so only in accordance with PTO guidelines stating that the requirement can be met by disclosing “sufficiently detailed, relevant identifying characteristics,” including “functional characteristics when coupled with a known or disclosed correlation between function and structure.” Univ. of Rochester v. G.D. Searle , 68 USPQ2d 1424, 1432 (DC WNY 2003). Therefore, only the above chemically structurally defined EXT1 shRNAs in Table 5 , the specific EXT1 guide RNA of Tokyo, and the shRNA sequences of Chen et al., but not the full breadth of the claim(s) meet the written description provision of 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA), first paragraph. The species specifically disclosed are not representative of the genus because the genus is highly variant. Applicant is reminded that Vas-Cath makes clear that the written description provision of 35 USC § 112 is severable from its enablement provision. (See page 1115.) 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. Claims 16- 2 2 and 25 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Tokyo ( EP3604502, Published 5 Feb 2020 ) , cited on an IDS. Regarding claim s 16 and 18 , Tokyo teaches methods for stably propagating virulent hand, foot and mouth disease viruses (paragraph 0007) and host cell s therefore that preferably do not express the EXT1 gene (paragraph s 0011 ,0035 ). Tokyo teaches a method for stably producing a virulent hand, foot and mouth disease virus comprising the steps of introducing genomic RNA of the virulent hand, foot and mouth disease virus into the host cell described above so as to obtain a cell producing the virus (paragraph s 0014 , 0041 ). Tokyo teaches introduction of the genomic RNA of the virus to the host cells by transfection (paragraph 0043). Tokyo teaches the “virulent hand, foot and mouth disease virus strain” is a population of multiple causative virus particles of hand, foot and mouth disease (paragraph 0050). Therefore, Tokyo teaches providing host cells that do not express the EXT1 gene and introducing genomic RNA of the hand, foot and mouth disease virus into the host cell to obtain a cell producing the virus particles. Regarding claim 17, Tokyo teaches the cell is preferably an RD cell (reading on a eukaryotic cell as this is a rhabdosarcoma cell) and teaches preparation of the RD cells by genome editing using guide RNA targeting EXT1 gene (paragraph s 0013 , 0072 ) . Regarding claim 19, Tokyo teaches EXT1 as a gene encoding an enzyme involved in the biosynthesis of heparan sulfate and that biosynthesis of heparin sulfate is inactivated due to a gene encoding the enzyme not being expressed ( paragraph 0035) and the expression of the gene involved in biosynthesis of heparan sulfate can be deleted by a method known in the art, such as knockout by gene editing using CRISPR/Cas9 or suppressed (knockdown) by gene silencing using siRNA or the like, and cites the GenBank Accession No. NM_000127 for human EXT1 gene (paragraph 0036). Regarding claims 20-22, Tokyo teaches preparing RD cells expressing no heparan sulfate by genome editing using CRISPR/Cas9, and the sequence of the guide RNA targeting the EXT1 gene was designed and shown below (paragraph 0072). Regarding claim 25, Tokyo teaches harvesting the virulent hand, foot and mouth disease virus propagated by culturing the cell obtained by step 1 (paragraph 0041), and that the harvesting can be by a heretofore known method, for example by repeated sonication or freeze-thaw cycling of the culture solution containing the virus-producing cells so as to destroy the virus-producing cells, followed by removal of cell debris by centrifugation, and may be further purified by polyethylene glycol precipitation or density gradient ultracentrifugation (paragraph 0047). 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. Claim s 22-24 are rejected under 35 U.S.C. 103 as being unpatentable over Tokyo as applied to claim s 16-2 2 and 25 above, and further in view of Chen et al. ( English translation of CN107058476 , Published 18 August 2017) , cited on an IDS . Claim Interpretation: Claim 22 was rejected in the 102(a)(1) rejection above for the species of guide RNA, and claim 22 is also included in this 103 rejection to reject an additional species of inhibitor recited in claim 22 (shRNA). The teachings of Tokyo as applicable to claims 16-22 and 25 h ave been described above. Tokyo does not teach that the inhibitor of EXT1 expression is an shRNA , or an shRNA represented by SEQ ID NO: 2. However, before the effective filing date , Chen et al. also taught the coding sequence of the EXT1 gene (NCBI Accession Number NM_000127) was known (page 6 of English translation) and t aught the construction of liver cancer cell lines that interfere with EXT1 expression using EXT1 shRNA sequences (page 8 of English translation). The EXT1 shRNA sequence 3-1 of Chen et al. shown below is 100% identical to the shRNA sequence of instant SEQ ID NO: 2. Below is the alignment of instant SEQ ID NO: 2 ( Qy ) with the EXT1 shRNA Sequence 3-1 of Chen et al. (Db): Chen et al. taught transfection of the EXT1 shRNA sequences into Hep3-B and Huh-7 cells, and that it successfully interfered with the expression of EXT1 in Hep3-B and Huh-7 liver cancer cell lines (page 9 and Fig. 4, shown below ). Therefore, it would have been obvious to one of ordinary skill in the art before the effective filing date, to have substituted the guide RNA targeting the EXT1 gene used in the method of Tokyo with the EXT1 shRNA of Sequence 3-1 of Chen et al. with a reasonable expectation of success. There would be a reasonable expectation of success because both Tokyo and Chen et al. teach polynucleotides that target the same gene, EXT1, and this would amount to simple substitution of one known element for another to obtain predictable results. One of ordinary skill in the art would have been motivated to use an EXT1 shRNA, specifically the sequence of EXT1 shRNA 3-1 of Chen et al. in the method of Tokyo as an alternative means to inhibit the expression of EXT1 as Chen et al. taught that the EXT1 shRNA sequences including sequence 3-1 successfully interfered with the expression of EXT1 in Hep3-B and Huh-7 liver cancer cell lines. Accordingly, the limitations of claims 22-24 would have been prima facie obvious to one of ordinary skill in the art before the effective filing date. Claims 26- 30,33 and 34 are rejected under 35 U.S.C. 103 as being unpatentable over Toky o ( EP3604502, Published 5 Feb 2020 ) , cited on an IDS . Regarding claims 26 and 28, Tokyo teaches methods for stably propagating virulent hand, foot and mouth disease viruses (paragraph 0007) and host cells therefore that preferably do not express the EXT1 gene (paragraphs 0011,0035). Tokyo teaches a method for stably producing a virulent hand, foot and mouth disease virus comprising the steps of introducing genomic RNA of the virulent hand, foot and mouth disease virus into the host cell described above so as to obtain a cell producing the virus (paragraphs 0014, 0041). Tokyo teaches introduction of the genomic RNA of the virus to the host cells by transfection (paragraph 0043). Tokyo teaches the “virulent hand, foot and mouth disease virus strain” is a population of multiple causative virus particles of hand, foot and mouth disease (paragraph 0050). Therefore, Tokyo teaches providing host cells that do not express the EXT1 gene and introducing genomic RNA of the hand, foot and mouth disease virus into the host cell to obtain a cell producing the virus particles. Tokyo does not teach the specific recited order of the steps of transfecting the cell population with an oligonucleotide encoding the biological entity and then inhibiting EXT1 expression in the cell by using an inhibitor of EXT1 expression and/or activity. However, it would have been obvious to one of ordinary skill in the art to change the order in the production method as taught by Tokyo to provide a cell population, transfect the cell population with the genomic RNA of the hand, foot and mouth disease virus into the host cell, and then a step of inhibiting EXT1 expression in the cell using an EXT1 inhibitor to arrive at claim 26 with a reasonable expectation of success. See MPEP 2144.04 IV. C. Changes in Sequence of Adding Ingredients, Ex parte Rubin, 128 USPQ 440 (Bd. App. 1959) (Prior art reference disclosing a process of making a laminated sheet wherein a base sheet is first coated with a metallic film and thereafter impregnated with a thermosetting material was held to render prima facie obvious claims directed to a process of making a laminated sheet by reversing the order of the prior art process steps.). See also In re Burhans , 154 F.2d 690, 69 USPQ 330 (CCPA 1946) (selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results); In re Gibson, 39 F.2d 975, 5 USPQ 230 (CCPA 1930) (Selection of any order of mixing ingredients is prima facie obvious.). As applicant has claimed two different orders of steps in claims 16 and 26 and both process arrive at the same result of producing a biological entity in a cell, there does not appear to be any criticality with the order of the steps since both orders are recited in the instant claims. Therefore, it would be obvious to perform the steps of transfecting a cell population with an oligonucleotide encoding the biological entity and inhibiting EXT1 expression the cell using an inhibitor of EXT1 expression and/or activity in any order. Regarding claim 27, Tokyo teaches the cell is preferably an RD cell and teaches preparation of the RD cells by genome editing using guide RNA targeting EXT1 gene (paragraphs 0013, 0072). Regarding claims 29-30, Tokyo teaches preparing RD cells expressing no heparan sulfate by genome editing using CRISPR/Cas9, and the sequence of the guide RNA targeting the EXT1 gene was designed and shown below (paragraph 0072). Regarding claims 33 and 34, Tokyo teaches a step of culturing the cell obtained by the step of introducing genomic RNA of the hand, foot and mouth disease virus into the host cell described above so as to propagate the virus, followed by harvesting the virulent hand, foot and mouth disease virus propagated by culturing the cell obtained by the culturing step (paragraph 0041) , and that the virus is released into the culture medium or accumulated inside the producing cells , and that the harvesting can be by a heretofore known method, for example by repeated sonication or freeze-thaw cycling of the culture solution containing the virus-producing cells so as to destroy the virus-producing cells, followed by removal of cell debris by centrifugation, and may be further purified by polyethylene glycol precipitation or density gradient ultracentrifugation (paragraph 0047). Accordingly, the limitations of claims 26-30,33 and 34 would have been prima facie obvious to one of ordinary skill in the art before the effective filing date. Claims 3 0-3 2 are rejected under 35 U.S.C. 103 as being unpatentable over Tokyo as applied to claims 26- 30,33 and 34 above, and further in view of Chen et al. (English translation of CN107058476 , Published 18 August 2017) , cited on an IDS . Claim Interpretation: Claim 30 was rejected in the 103 rejection above for the species of guide RNA, and claim 30 is also included in this 103 rejection to reject an additional species of inhibitor recited in claim 30 (shRNA). The teachings of Tokyo as applicable to claims 26-30,33 and 34 h ave been described above. Tokyo do es not teach that the inhibitor of EXT1 expression is an shRNA or an shRNA represented by SEQ ID NO: 2. However, before the effective filing date, Chen et al. also taught the coding sequence of the EXT1 gene (NCBI Accession Number NM_000127) was known (page 6 of English translation) and taught the construction of liver cancer cell lines that interfere with EXT1 expression using EXT1 shRNA sequences (page 8 of English translation). The EXT1 shRNA sequence 3-1 of Chen et al. shown below is 100% identical to the shRNA sequence of instant SEQ ID NO: 2. Below is the alignment of instant SEQ ID NO: 2 ( Qy ) with the EXT1 shRNA Sequence 3-1 of Chen et al. (Db): Chen et al. t aught transfection of the EXT1 shRNA sequences into Hep3-B and Huh-7 cells, and that it successfully interfered with the expression of EXT1 in Hep3-B and Huh-7 liver cancer cell lines (page 9 and Fig. 4, shown below). Therefore, it would have been obvious to one of ordinary skill in the art before the effective filing date, to have substituted the guide RNA targeting the EXT1 gene used in the method of Tokyo with the EXT1 shRNA of Sequence 3-1 of Chen et al. with a reasonable expectation of success. There would be a reasonable expectation of success because both Tokyo and Chen et al. teach polynucleotides that target the same gene, EXT1, and this would amount to simple substitution of one known element for another to obtain predictable results. One of ordinary skill in the art would have been motivated to use an EXT1 shRNA, specifically the sequence of EXT1 shRNA 3-1 of Chen et al. in the method of Tokyo as an alternative means to inhibit the expression of EXT1 as Chen et al. taught that the EXT1 shRNA sequences including sequence 3-1 successfully interfered with the expression of EXT1 in Hep3-B and Huh-7 liver cancer cell lines. Accordingly, the limitations of claims 3 0-3 2 would have been prima facie obvious to one of ordinary skill in the art before the effective filing date. Conclusion Claims 16-34 are rejected. Any inquiry concerning this communication or earlier communications from the examiner should be directed to FILLIN "Examiner name" \* MERGEFORMAT STEPHANIE L SULLIVAN whose telephone number is FILLIN "Phone number" \* MERGEFORMAT (703)756-4671 . The examiner can normally be reached FILLIN "Work Schedule?" \* MERGEFORMAT Monday-Friday, 7:30-3:30 EST. 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. 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If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000. /STEPHANIE L SULLIVAN/ Examiner, Art Unit 1635 /ABIGAIL VANHORN/ Primary Examiner, Art Unit 1636