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 .
Response to Amendment/Status of Claims
Receipt of Arguments/Remarks filed on 07/01/2026 is acknowledged. Claims 4 and 5 were cancelled. Claims 1-3,17,21 and 34 were amended. Claims 68-75 are new. Claims 1-3,6-9,11-13,17,18,20,21,31,34,35 and 66-75 are pending and under examination.
Priority
This application is a 371 of PCT/US2022/019012 filed 03/04/2022, which claims benefit of 63/157,498, filed 03/05/2021 and claims benefit of 63/157,497, filed 03/05/2021 as reflected by the most recent filing receipt.
Withdrawn Rejections
Applicant’s arguments and amendments, see page 9, filed 07/01/2026, with respect to the objection to FIG. 7H have been fully considered and are persuasive due to providing a replacement figure having improved clarity by increasing the font size. The objection to FIG. 7H has been withdrawn.
Applicant’s arguments and amendments, see page 9, filed 07/01/2026, with respect to the 35 U.S.C. 112(b) rejection of claims 2 and 34 have been fully considered and are persuasive due to the amendments to claims 2 and 34 correcting the issues of indefiniteness. The 35 U.S.C. 112(b) rejection of claims 2 and 34 has been withdrawn.
Applicant’s arguments and amendments, see pages 9-14, filed 07/01/2026, with respect to the rejection(s) of claim(s) 1-3,8,34,66 and 67 under 35 U.S.C 102(a)(1) as anticipated by Church, claims 9,11,17,18,20 and 21 under 35 U.S.C. 103 as unpatentable over Church, claims 4-7,31 and 35 as unpatentable over Church and further in view of Harvard, claim 12 as unpatentable over Church and further in view of Szybalski, and claim 13 as unpatentable over Church and further in view of Jones, have been fully considered and are persuasive due to the amendments made to the claims in which the cited references do not teach all of the limitations of the claims as amended. Therefore, the rejection has been withdrawn. However, upon further consideration, a new ground(s) of rejection is made in view of the amendments to claims 1-3,17,21 and 34 as well as adding new claims 68-75. See the new 103 rejections below which include a new reference to teach the limitations of the claim amendments. While the 103 rejections still use Church and Harvard, Lin et al. (WO 2020201434, Published 08 Oct 2020) is newly applied to teach the limitations of the claims as amended, and therefore Applicant’s arguments regarding Church and Harvard are not found persuasive because Lin et al. is used to teach the remaining claim limitations of the amended claims.
Claim Objections
Claim 1 is objected to because of the following informalities: step (b) and step (e) each recite “recipient oligonuclide” and therefore “oligonuclide” is a mis-spelling and should be corrected to “oligonucleotide”. Appropriate correction is required.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 8,9,21,72 and 74 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 8 depends on claim 1 and recites the limitation “wherein the first, second or subsequent donor plasmid” in lines 1-2. There is insufficient antecedent basis for this limitation in the claim because there is a first and second donor plasmid in claim 1 but no recitation of a subsequent donor plasmid in claim 1 so lacks antecedent basis for the subsequent donor plasmid.
Claim 9 depends on claim 1 and recites the limitation “the first, second, third or subsequent oligonucleotide” in lines 2-3. There is insufficient antecedent basis for this limitation in the claim because while there is a first and second oligonucleotide recited in claim 1, there is no recitation of a third oligonucleotide or a subsequent oligonucleotide in claim 1 so lacks antecedent basis.
Claim 21 depends on claim 1 and recites “the first, second or subsequent homologous recombination (HR) regions and their corresponding HR region on the recipient oligonucleotide….” in lines 1-3. There is insufficient antecedent basis for the limitation of subsequent homologous recombination regions in the claim because there is no recitation of “subsequent” homologous recombination regions in claim 1 and therefore lacks antecedent basis.
Claim 72 depends on claim 1 and recites “the counter selectable marker”. There is insufficient antecedent basis for this limitation in the claim because there is no prior recitation of “the counter selectable marker” and claim 1 only recites a selectable marker.
Claim 74 depends on claim 1 and recites “the one or more homologous DNA repair genes”. There is insufficient antecedent basis for this limitation in the claim because there is no prior recitation of “one or more homologous DNA repair genes” in claim 1.
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claims 8,11,66 and 71 are rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends.
Claim 8 recites, “The method of claim 1, wherein the first, second or subsequent donor plasmid comprises a selectable marker selecting for integration of the first oligonucleotide, second oligonucleotide or subsequent oligonucleotide into the recipient oligonucleotide”. Claim 1 has been amended to recite that each of the first and second donor plasmids comprises a selectable marker, and therefore claim 8 does not recite a further limitation of the subject matter claimed.
Claim 11 recites, “wherein the donor plasmid comprises a conditional replication origin”. Amended claim 1 now recites “wherein each donor plasmid comprises an origin of transfer (oriT) and a conditional replication origin”. Therefore, claim 11 does not recite a further limitation as claim 1 already recites the donor plasmid comprises a conditional replication origin.
Claim 66 recites “wherein HR2.1 and HR2.2 flank a non-homologous region comprising one (C2) or two endonuclease sites (C2.1, C2.2);HR3 and HR4 flank a non-homologous region comprising one (C4) or two endonuclease sites (C4.1, C4.2); and/or HR6.1 and HR6.2 flank a non-homologous region comprising one (C7) or two endonuclease sites (C7.1, C7.2)”. However, amended claim 1 now recites wherein HR2.1 and HR2.2 flank a non-homologous region (NHR) comprising two endonuclease sites (C2.1, C2.2); HR3 and HR4 flank a non-homologous region comprising two endonuclease sites (C4.1, C4.2); and wherein HR6.1 and HR6.2 flank a non-homologous region (NHR) comprising two endonuclease sites (C7.1, C7.2) . Therefore, claim 66 fails to recite a further limitation of claim 1.
Claim 71 recites “wherein the selectable marker is within a non-homologous region between HR2.1 and HR2.2 and/or between HR6.1 and HR6.2, and/or between subsequent HR regions. This fails to recite a further limitation of claim 1, because amended claim 1 recites, “wherein HR2.1 and HR2.2 flank a non-homologous region (NHR) comprising two endonuclease sites (C2.1, C2.2) flanking a selectable marker”…
“wherein HR6.1 and HR6.2 flank a non-homologous region (NHR) comprising two endonuclease sites (C7.1, C7.2) flanking a selectable marker”.
Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
Claim Rejections - 35 USC § 112
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
New Matter Rejection
Claims 1-3,6-9,11-13,17,18,20,21,31,34,35 and 66-75 introduce new matter as claim 1 has been amended to recite the limitations: “wherein HR2.1 and HR2.2 flank a non-homologous region (NHR) comprising two endonuclease sites (C2.1, C2.2) flanking a selectable marker”; “and HR3 and H4 flank a non-homologous region comprising two endonuclease sites (C4.1, C4.2) flanking a selectable maker; (b) subjecting the first donor plasmid and recipient oligonuclide to endonuclease digestion with a first endonuclease to cleave the first donor plasmid at C1 and C3 and the recipient oligonucleotide at C4.1 and C4.2, thereby producing a first donor cassette comprising homologous recombination regions HR1 and HR2.2 at each termini and a recipient oligonucleotide having compatible recombination regions HR3 and HR4; (c) subjecting the first donor cassette and the recipient oligonucleotide to conditions to recombine the first donor cassette into the recipient oligonucleotide by homologous recombination”; “wherein HR6.1 and HR6.2 flank a non-homologous region (NHR) comprising two endonuclease sites (C7.1, C7.2) flanking a selectable marker;(e) subjecting the second donor plasmid and recipient oligonucleotide to endonuclease digestion with a second endonuclease to cleave the second donor plasmid at C5 and C6 and the recipient oligonucleotide at C2.1 and C2.2, thereby producing a second donor cassette comprising homologous recombination regions HR5 and HR6.2 at each termini and a recipient oligonucleotide having compatible recombination regions HR2.1 and HR4; (f) subjecting the second donor cassette and the recombined recipient oligonucleotide to conditions to recombine the first donor fragment and the recombined recipient oligonucleotide in the recipient cell by homologous recombination”; “wherein each donor plasmid comprises an origin of transfer (oriT) and a conditional replication origin; wherein an oligonucleotide encoding a guide RNA or a first endonuclease targeting the first, third, and/or fourth endonuclease sites is present on the first donor plasmid and/or is present in the recipient cell, and wherein an oligonucleotide encoding a guide RNA or a second endonuclease targeting the second, fifth, and/or sixth endonuclease site is present on the second donor plasmid and/or is present in the recipient cell”.
From MPEP 2163.06: “Applicant should therefore specifically point out the support for any amendments made to the disclosure.” Applicant has not directed the Examiner to the support in the specification for the amendments. Numerous amendments have been made to claim 1 throughout the claim and other than stating that support for the amendments is found in the specification and claims as originally filed, Applicant did not point to any specific areas to support these numerous amendments to claim 1.
It is not clear to the Examiner that the method in claim 1 as now claimed has written support in the specification. The Examiner does not see a method disclosed with all of these steps (a)-(f) as recited in the amended claims. While there may be support for some of the elements and parts of the method, the Examiner does not see support for all of the limitations. For Example, there does not appear to be any specific recitation of an “endonuclease digestion step” recited in steps (b) and (e). In addition, regarding the “first donor cassette” recited in claim 1 in new step (b), there is only one mention of “donor cassette” in whole application (paragraph 0298): “Construction of swapping cassettes: A swapping cassette is defined as the stretch of DNA on the donor and recipient plasmids that participates in a DNA swap: the cassette on the recipient plasmid is replaced by the cassette originally found on the donor plasmid via homologous recombination. In order to recursively select for cassette swapping in vivo, each cassette is engineered to contain both a selectable and counter-selectable marker. A selectable marker in the donor cassette and a counter-selectable marker in the recipient cassette is needed in every round”. However this does not say what step (b) recites in claim 1.
Therefore, it is the Examiner' s position that the disclosure does not reasonably convey that the inventor had possession of the subject matter of the amendment at the time of filing of the instant application.
Claims 2,3,6-9,11-13,17,18,20,21,31,34,35 and 66-75 are included in the new matter rejection because they depend from claim 1 which is rejected as containing new matter and therefore also have the same issues.
Claim Rejections - 35 USC § 103
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Claims 1-3,6-9,11,17,18,20,21,31,34,35,66-68,70-72 and 74 are rejected under 35 U.S.C. 103 as being unpatentable over Church (US 20070004041, Published 4 Jan 2007), cited on an IDS as evidenced by Kvitko et al. (BMC Research Notes, 2012, 5:157), in view of Lin et al. (WO 2020201434, Published 08 Oct 2020) and Harvard (US 20200392538, Published 17 Dec 2020), cited on an IDS.
Claim Interpretation: Regarding the homologous DNA repair genes and recombination-mediated genetic engineering genes of claims 17,18 and 74, the instant specification discloses that the donor cell or recipient cell of the methods described herein includes an oligonucleotide encoding one or more homologous DNA repair genes, such as RecA, and in embodiments the homologous DNA repair gene is RecA, and in embodiments the homologous DNA repair genes are the recombineering genes Red alpha, Red beta and Red gamma (paragraph 0120). The instant specification discloses that in embodiments, the recombineering genes are lambda red genes; the recombination-mediated genetic engineering genes are Red alpha, Red beta, and Red gamma (paragraph 0148). Therefore, art teaching RecA reads on a homologous DNA repair gene for claim 17, and art teaching Red alpha, Red beta or Red gamma reads on recombination-mediated genetic engineering genes for claim 18.
Regarding claims 1,66,71 and 72, Church teaches a method for hierarchical assembly of very large, including genome sized, nucleic acid products (paragraph 0123) and the polynucleotide constructs assemble in a desired manner by integrating into the host cell genome by homologous recombination (paragraph 0018) and the polynucleotide constructs may be contained on an extrachromosomal plasmid (paragraph 0019). Church teaches the hierarchical assembly process is illustrated with reference to Figs. 2-4 (paragraph 0129).
Church teaches for each pairwise combination, the donor and recipient strains are mixed and the donor cells transfer their DNA to the recipient cells by conjugation (paragraphs 0068, 0129). Church teaches that Fig. 4 illustrates an embodiment of a hierarchical assembly method, which involves two sets of starting polynucleotide constructs each having four types of components including selected genes (illustrated as 2 and 3 in Fig. 4), meganuclease sites (illustrated as 4 and 5 in Fig. 4), conjugative transfer sites (oriT sites, illustrated as 6 and 7 in Fig. 4), and origins of replication (illustrated as 8 and 9 in Fig. 4) and the polynucleotide constructs are introduced into cells, the cells are then mixed pairwise and in each case the cell containing the lower polynucleotide construct of the pair (donor) is transferred into the cell containing the upper polynucleotide construct of the pair (recipient), and the cell containing the upper polynucleotide construct contains a meganuclease that recognizes the meganuclease cleavage sites of the incoming (lower) polynucleotide construct, and cleavage by the meganuclease stimulates homologous recombination between the upper and lower polynucleotide constructs (paragraph 0136).
Regarding the first donor plasmid and recipient oligonucleotide, Church teaches starting materials include a cell and polynucleotide constructs that together comprise the sequence of the modified genome (hatched; labeled A,B,C and D) in which polynucleotide construct A is shown as containing a first meganuclease site at the beginning of the construct and a second meganuclease site at the end of the construct (Fig 3A and paragraphs 0132, 0139), and Fig. 4 shows a donor cell comprising first and second meganuclease sites (illustrated as 4 and 5 in Fig. 4) a first recombination site b (equivalent to instant HR1) and a second recombination site yz (equivalent to instant HR 2.1 and 2.2) flanking a sequence of interest (equivalent to instant oligo 1), and the recipient oligonucleotide comprises a third homologous recombination region (HR3) homologous to HR1 and a fourth homologous recombination region (HR4) homologous to HR2.2 as Fig. 4 teaches in Round 1 a recipient comprising a first recombination site b (equivalent to instant HR3) and a second recombination site yz (equivalent to instant HR4) flanking a sequence of interest 3 (equivalent to instant oligo 1) thereby providing following the homologous recombination of HR1 with HR3 and HR2.2 with HR4, a first recombined recipient oligonucleotide comprising the first DNA element fragment.
Church teaches assembly into larger polynucleotide constructs, or whole genome replacement is achieved by repeated rounds of conjugation and integration (Fig. 3C and D, paragraph 0133) and teaches contacting the recipient cell with a second donor cell under conditions to transfer the second donor plasmid from the second donor cell to the first recipient cell by conjugation (Fig. 4). Church teaches that the product produced by introduction of the polynucleotide constructs are a plurality of cells comprising a single integrated polynucleotide construct replacing the wild-type sequence at that location (Fig 3B, paragraph 0132). Church teaches providing a plurality of polynucleotide constructs comprising a sequence encoding a first selectable marker and a portion of the plurality of polynucleotide constructs comprise a sequence encoding a second selectable marker (Fig. 3 and paragraphs 0012-0014,0024,0132), and the desired product may be selected using the appropriate selectable marker (see FIG. 3C, paragraph 0133).
Church teaches that Fig. 3 discloses a second donor plasmid comprising a first homologous region flanking AB (equivalent to instant HR5) and a second homologous region flanking AB (equivalent to instant HR6), and the final modified genome produced by further rounds of conjugation and homologous recombination in which recombination between homologous region flanking AB with homologous region flanking CD forms a DNA assembly (Fig. 3D, paragraph 0099). Figure 4 again shows the second donor plasmids (Round 2) as comprising a first and second endonuclease site (meganuclease sites illustrated as 4 and 5) and the recombination regions.
PNG
media_image1.png
731
580
media_image1.png
Greyscale
PNG
media_image2.png
722
994
media_image2.png
Greyscale
Regarding the conditional replication origin in claims 1, Church teaches the upper cell supports replication of the upper polynucleotide construct but not does not support replication of the lower (incoming) polynucleotide construct. This may be achieved using different conditional origins of replication such as, for example, IncX R6K oriγ (dependent on pir protein) or IncPα ori V (dependent on the trfA protein) for the polynucleotide constructs contained in the donor and recipient cells (paragraph 0137). Conditional origins of replication are origins that require the presence or expression of a trans-acting factor in the host cell for replication. A variety of conditional origins of replication functional in prokaryotic hosts (e.g., E. coli) are known to the art. Exemplary conditional origins of replication that may be used in accordance with the hierarchical assembly methods and genome excision methods described herein include, for example, the R6Kγ origin (paragraph 0149).
Church teach for purposes of illustration only, FIG. 4 utilized a combination of two selectable markers, two meganuclease cleavage sites, two conjugative transfer elements and two origins of replication but it may be desirable to use 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more, different selectable markers, meganuclease cleavage sites, conjugative transfer elements, and/or origins of replication (paragraph 0139).
While Church suggests that more than two meganuclease cleavage sites can be used, Church does not teach the donor plasmids and recipient oligonucleotides contain two endonuclease sites flanking a selectable marker as recited in claim 1. Church does not teach an oligonucleotide encoding a guide RNA or a first endonuclease targeting the first, third and/or fourth endonuclease site present on the first donor plasmid, or an oligonucleotide encoding a guide RNA or a second endonuclease targeting the second, fifth and/or sixth endonuclease site present on the second donor plasmid.
Before the effective filing date, Lin et al. taught DNA assembly methods and that an ongoing struggle is to achieve a balance between reducing sequence design constraints such as forbidden sequence in assembled DNA and unwanted scar sequence introduced during the assembly process, and improving assembly modularity and part reusability (page 1, lines 4,14-17). Lin et al. taught the initial type IIS restriction enzyme-based DNA assembly system was named Golden Gate assembly. In Golden Gate assembly system, insert DNA fragments to be assembled within insert plasmids are flanked by recognition sites for a type IIS restriction enzyme that cuts outside the recognition sequence and generates arbitrary adhesive ends with sequences independent of the enzyme recognition sequence. The insert plasmids for Golden Gate assembly contain inserts flanked by type IIS restriction sites within the insert vector backbone facing the inserts, so that once the inserts are released from the plasmid by the type IIS restriction enzyme, they do not contain the restriction sites. The assembly vector for Golden Gate assembly contains a negative selection marker flanked by type IIS restriction enzyme sites located in the selection marker facing the vector, so that once the selection marker is released from the vector, the vector backbone does not contain the type IIS restriction sites. These specially designed plasmids allow simple one-pot assembly of multiple DNA fragments by mixing insert plasmids and vectors together with a type IIS restriction enzyme and DNA ligase (page 1, lines 19-33).
Lin et al. taught, advantageously, the presence of the sequence-specific DNA-binding protein protects the type IIS restriction enzyme recognition sequence from methylation, and effectively switches ON the type IIS restriction recognition sequence such that it can be cut by the type IIS restriction enzyme, and where the sequence-specific DNA- binding protein is not present, the DNA methylase may bind to and methylate the type IIS restriction recognition sequence, such that it is protected by methylation and effectively switches OFF the type IIS restriction recognition sequence such that the type IIS restriction enzyme cannot cut the nucleic acid. Described herein is a new method for type IIS restriction enzyme-based DNA assembly that overcomes the problem of sequence constraint and so eliminates the requirement to remove internal type IIS restriction sites from DNA parts to assemble. The method, herein termed “universal assembly”, is based on the methylation protection approach, whereby a DNA methylase is used to methylate and so block any internal restriction sites for the type IIS restriction enzyme in any DNA fragment to be assembled. In parallel, a sequence-specific DNA binding protein, such as a deactivated and programmable CRISPR Cas9, or other sequence-specific DNA binding protein, can be used to bind near to and prevent methylation of particular type IIS restriction sites of methylation-protectable restriction elements, which can be positioned in DNA vectors to flank DNA inserts or fragments to be excised. This advantageously provides control of the restriction digestion and release of the DNA insert/fragment during the assembly process (pages 4-5).
Harvard also teaches methods for assembly of synthetic genomes using nuclease assisted homologous recombination which enable scarless and iterative replacement of wild-type DNA with large (at least 50 kb) synthetic DNA segments at desired genomic loci (paragraph 0004). Harvard recites a method of introducing into a parental cell a donor DNA segment flanked by first homology sequences, wherein the parental cell comprises a selectable marker gene and flanked by second homology sequences homologous to the first homology sequences, and an inducible recombineering system, introducing into the parental cell a RNA-guide nuclease or nucleic acid encoding the RNA-guide nuclease and at least one nucleic acid encoding at least one guide RNA targeting the selectable marker gene and inducing expression of the inducible recombineering system (claim 2), and recites the RNA-guided nuclease is Cas9 nuclease (claims 15-16) (an RNA-guided endonuclease). Harvard teaches that gRNA targets cutting of the selectable marker gene in the genome of the parent cell to initiate and enhances recombination (paragraph 0007).
Harvard teaches the parental cells are engineered to carry an inducible sequence-specific nuclease (e.g., genomically), thus, nuclease activity may be induced before, during or after induction of expression of the recombineering system (paragraph 0011) and recites the expression of the nucleic acid encoding the RNA-guided nuclease is inducible (claims 67-68). Harvard teaches a RNA-guided nuclease (e.g., Cas9) and guide RNA (gRNA) targeting the genomic loci for integration are introduced in the cells (e.g., by plasmid transformation), before, during or after induction of expression of the recombineering system and in other embodiments, the parental cells are engineered to carry a RNA-guided nuclease (e.g., genomically), thus, only the gRNA targeting the genomic loci for integration is introduced before, during or after induction of expression of the recombineering system (paragraph 0012).
It would have been obvious to one of ordinary skill in the art before the effective filing date, to modify the DNA assembly method of Church with the teachings of Lin et al. regarding the assembly vector for Golden Gate assembly containing a selection marker flanked by type IIS restriction enzyme sites and Harvard regarding the DNA assembly method involving gRNA and Cas9 with a reasonable expectation of success. There would be a reasonable expectation of success as Church, Lin et al. and Harvard pertain to DNA assembly using nuclease assisted homologous recombination and would amount to combining prior art elements according to known methods to yield predictable results. One of ordinary skill in the art would have been motivated to do so because Lin et a. taught for DNA assembly methods the need to achieve a balance between reducing sequence design constraints such as forbidden sequence in assembled DNA and unwanted scar sequence introduced during the assembly process, and improving assembly modularity and part reusability (page 1, lines 4,14-17), the assembly vector for Golden Gate assembly contains a negative selection marker flanked by type IIS restriction enzyme sites located in the selection marker facing the vector, so that once the selection marker is released from the vector, the vector backbone does not contain the type IIS restriction sites. These specially designed plasmids allow simple one-pot assembly of multiple DNA fragments by mixing insert plasmids and vectors together with a type IIS restriction enzyme and DNA ligase (page 1, lines 19-33). Lin et al. also taught a new method for type IIS restriction enzyme-based DNA assembly that overcomes the problem of sequence constraint and so eliminates the requirement to remove internal type IIS restriction sites from DNA parts to assemble, called “universal assembly”, based on the methylation protection approach, whereby a DNA methylase is used to methylate and so block any internal restriction sites for the type IIS restriction enzyme in any DNA fragment to be assembled. In parallel, a sequence-specific DNA binding protein, such as a deactivated and programmable CRISPR Cas9, or other sequence-specific DNA binding protein, can be used to bind near to and prevent methylation of particular type IIS restriction sites of methylation-protectable restriction elements, which can be positioned in DNA vectors to flank DNA inserts or fragments to be excised. This advantageously provides control of the restriction digestion and release of the DNA insert/fragment during the assembly process (pages 4-5). One of ordinary skill in the art would have been motivated to modify the method of Church with the teachings of Harvard because Harvard teach a method of introducing into a parental cell a donor DNA segment flanked by first homology sequences, wherein the parental cell comprises a selectable marker gene and flanked by second homology sequences homologous to the first homology sequences, and an inducible recombineering system, introducing into the parental cell a RNA-guide nuclease or nucleic acid encoding the RNA-guide nuclease and at least one nucleic acid encoding at least one guide RNA targeting the selectable marker gene and inducing expression of the inducible recombineering system (claim 2), and recites the RNA-guided nuclease is Cas9 nuclease (claims 15-16) (an RNA-guided endonuclease). Harvard teaches that gRNA targets cutting of the selectable marker gene in the genome of the parent cell to initiate and enhances recombination (paragraph 0007), and the parental cells are engineered to carry an inducible sequence-specific nuclease (e.g., genomically), thus, nuclease activity may be induced before, during or after induction of expression of the recombineering system (paragraph 0011). Modifying Church with the teachings of Lin et al. and Harvard et al. regarding the arrangement and structures of the donor plasmids and recipient oligonucleotides and the endonuclease cut sites as well as selectable markers would result in the functional limitations regarding cleaving at specific sites and would result in the recited structures.
Regarding claims 2 and 3, Church teaches the process is repeated using further rounds of pairwise mixing, conditional cleavage of the incoming polynucleotide construct at the meganuclease site, selection of the incoming selectable marker and facilitated loss of the unwanted portions of the incoming polynucleotide construct using a conditional original of replication and/or negative selectable marker (see e.g. Rounds 2-4 in Fig. 4) until the desired product is achieved (paragraph 0138). Figure 4 shows repeated iterations of third and subsequent donor cells for conjugation with the recipient cell and discloses a plurality of donor cells for each of the first and second rounds.
Regarding claims 6 and 7, Church does not teach expression of the first and/or the second endonuclease is inducible and the method comprises inducing expression of the first and/or second endonuclease, or that the first and/or second endonuclease is an RNA-guided endonuclease.
Harvard teaches the parental cells are engineered to carry an inducible sequence-specific nuclease (e.g., genomically), thus, nuclease activity may be induced before, during or after induction of expression of the recombineering system (paragraph 0011) and recites the expression of the nucleic acid encoding the RNA-guided nuclease is inducible (claims 67-68). Harvard teaches a RNA-guided nuclease (e.g., Cas9) and guide RNA (gRNA) targeting the genomic loci for integration are introduced in the cells (e.g., by plasmid transformation), before, during or after induction of expression of the recombineering system and in other embodiments, the parental cells are engineered to carry a RNA-guided nuclease (e.g., genomically), thus, only the gRNA targeting the genomic loci for integration is introduced before, during or after induction of expression of the recombineering system (paragraph 0012).
It would have been obvious to one of ordinary skill in the art before the effective filing date, to modify the DNA assembly method of Church with the teachings of Lin et al. regarding the assembly vector for Golden Gate assembly containing a selection marker flanked by type IIS restriction enzyme sites and teachings of Harvard regarding a first and/or second endonuclease is that is inducible, and wherein the endonuclease is an RNA-guided endonuclease (Cas9). One of ordinary skill in the art would be motivated to provide an inducible first and/or second endonuclease and to induce expression of the first and/or second endonuclease because Harvard recites the expression of the nucleic acid encoding the RNA-guided nuclease is inducible (claims 67-68) and teaches the parental cells are engineered to carry an inducible sequence-specific nuclease (e.g., genomically), thus, nuclease activity may be induced before, during or after induction of expression of the recombineering system (paragraph 0011). One of ordinary skill in the art would be motivated to provide an RNA-guided endonuclease as the first and/or second endonuclease because Harvard teaches a RNA-guided nuclease (e.g., Cas9) and guide RNA (gRNA) targeting the genomic loci for integration are introduced in the cells (e.g., by plasmid transformation), before, during or after induction of expression of the recombineering system and in other embodiments and the parental cells are engineered to carry a RNA-guided nuclease (e.g., genomically) (paragraph 0012).
Regarding claim 8, Church teaches providing a plurality of polynucleotide constructs comprising a sequence encoding a first selectable marker and a portion of the plurality of polynucleotide constructs comprise a sequence encoding a second selectable marker (Fig. 3 and paragraphs 0012-0014,0024,0132), and the desired product may be selected using the appropriate selectable marker (see FIG. 3C, paragraph 0133).
Regarding claim 9, Church teaches in certain embodiments, negative or counter selectable markers could be placed on the polynucleotide constructs to facilitate loss of the unwanted portions of the donor polynucleotide construct that are transferred to the recipient strain. The negative selectable marker may be incorporated into the polynucleotide construct outside of the region that homologously recombines with the recipient genome such that portions of the polynucleotide construct not incorporated into the genome may be removed by negative selective pressure and/or cells which have incorporated undesired regions of the polynucleotide construct into the recipient genome may be removed using negative selective pressure (paragraph 0137), and that polynucleotide constructs may comprise both selectable and counter selectable markers (paragraph 0143).
Regarding claim 11, the conditional replication origin taught by Church has been described above pertaining to claim 1.
Regarding claims 17 and 74, Church teaches the frequency of homologous recombination in prokaryotes is significantly enhanced by the presence of recombinase activities and several purified proteins catalyze homologous pairing and/or strand exchange in vitro, including: E. coli recA protein (the inducible homologous DNA repair gene of claims 17 and 74), and recombinases, like the recA protein of E. coli are proteins which promote strand pairing and exchange. Church teaches the most studied recombinase to date has been the recA recombinase of E. coli, which is involved in homology search and strand exchange reactions (paragraph 0189). Church teaches RecA is required for induction of the SOS repair response, DNA repair, and efficient genetic recombination in E. coli, and that RecA can catalyze homologous pairing of a linear duplex DNA and a homologous single strand DNA in vitro. In contrast to site-specific recombinases, proteins like recA which are involved in general recombination recognize and promote pairing of DNA structures on the basis of shared homology, as has been shown by several in vitro experiments (paragraph 0189).
Regarding claim 18, Church teaches the polynucleotide constructs assemble in a desired manner by integrating into the host cell genome by homologous recombination, site-specific recombination, or combinations thereof, and when assembly involves homologous recombination, the host cells may express a recombinase such as, for example, recE and recT from E. coli or the Redα and Redβ proteins from lambda (paragraph 0018). Church teaches each polynucleotide construct is separately introduced into a cell, for example, constructs then integrate into the host cell genome by homologous recombination, and that it may be desirable to utilize a host cell that is overexpressing a recombinase and/or comprises a recombinase under the control of an inducible/repressible promoter. Exemplary recombinase systems include, for example, RedE and RecT proteins (from E. coli) or Redα, Redβ and Gam proteins (from lambda) (paragraph 0132).
Regarding claims 20,21 and 68, Church teaches the plurality of polynucleotide segments are each at least about 50 kilobases in length, 100 kilobases in length, or longer (paragraph 0047) and embodiments wherein at least one terminal sequence of each polynucleotide construct is homologous with the terminal sequences of another polynucleotide construct. Such homologous terminal regions may be at least about 20, 50, or more nucleotides in length (paragraph 0020). It is generally noted that differences in a range of oligonucleotide length do not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such length is critical. "[W]here the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation." In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955). Given that applicant did not point out the criticality of the length of the assembled DNA element of the invention, it is concluded that the normal desire of scientists or artisans to improve upon what is already generally known would provide the motivation to determine where in a disclosed set of ranges is the optimum length NOTE: MPEP 2144.05.
Regarding claims 31 and 70, Church does not teach the method using two or more recipient oligonucleotides having compatible homologous recombination regions to construct a DNA library or that the method generates a combinatorial library comprising a plurality of different assembled DNA elements.
Harvard teaches the methods of the present disclosure are used for multiplexed integration of large-size DNA libraries, for example recoded segment libraries.
It would have been obvious to one of ordinary skill in the art before the effective filing date, to modify the method of Church to use two or more recipient oligonucleotides to construct a DNA library or to generate a combinatorial library comprising a plurality of different assembled DNA elements based on the teachings of Harvard with a reasonable expectation of success. One of ordinary skill in the art would have been motivated to do so based on the teachings of Harvard suggesting that the methods of Harvard can be used for multiplexed integration of large-size DNA libraries, including recoded segment libraries.
Regarding claims 34 and 67, Church teaches the assembly methods described herein require polynucleotide constructs for assembly of large product nucleic acids, such as, for example, a modified, partially synthetic, or fully synthetic genome. The sequences obtained from such sources may then be modified using standard molecular biology and/or recombinant DNA technology to produce polynucleotide constructs having desired modifications for reintroduction into, or construction of, a large product nucleic acid, including a modified, partially synthetic or fully synthetic genome (paragraph 0155). Therefore, Church teaches the method is used to combine genetic regions of genes.
Regarding claim 35, Church does not teach the first and second oligonucleotides comprising the first and second DNA element fragments are inserted into the first and second donor plasmids prior to steps (a) and (b).
Harvard teaches methods and compositions (e.g., cells, genetic constructs, and kits) for targeted scarless integration of large DNA segments (e.g., at least 50 kb) from a donor into a receiver (parental) strain genome, and each assembly cycle (integration of a single segment) of these methods for genome assembly can be iterated for integration of multiple donor DNA segments in a sequential or parallel manner. Harvard teaches donor DNA segments may be assembled on a plasmid from synthetic DNA segments and the donor DNA is then introduced into the parental cells (paragraph 0010). Therefore, Harvard teaches insertion of donor DNA segments into the plasmid prior to the donor DNA segments being introducing into the parental cells (donor cells).
It would have been obvious to one of ordinary skill in the art before the effective filing date, to modify the method of Church to include a step of inserting the first and second oligonucleotides comprising the first and second DNA element fragments into the first and second donor fragments prior to the steps of contacting the first donor cells comprising the donor plasmids with the recipient cells based on the teachings of Harvard with a reasonable expectation of success. One of ordinary skill in the art would have been motivated to do so because Harvard teaches methods for genome assembly can be iterated for integration of multiple donor DNA segments in a sequential or parallel manner and that donor DNA segments may be assembled on a plasmid from synthetic DNA segments and the donor DNA is then introduced into the parental cells (paragraph 0010).
Regarding claim 73, the teachings of Church regarding the conditional origin of replication have been described above pertaining to claim 1. Church taught conditional origins of replication include IncX R6K oriγ (dependent on pir protein) or IncPα ori V (dependent on the trfA protein) for the polynucleotide constructs contained in the donor and recipient cells (paragraph 0137) Conditional origins of replication are origins that require the presence or expression of a trans-acting factor in the host cell for replication.
As evidenced by Kvitko et al., the γ origin of replication of the broad-host-range plasmid R6K (oriR6Kγ) has been used to construct conditionally replicative cloning and transposon delivery vectors too numerous to cite them all and replication of these vectors requires the π protein encoded by the pir gene which on R6K is located next to the γ origin of replication. Therefore, Church et al. teach a conditional replication origin which is dependent on the presence of an oligonucleotide (pir protein encoded by the pir gene).
Accordingly, the limitations of claims 1-3,6-9,11,17,18,20,21,31,34,35,66-68 and 70-74 would have been prima facie obvious to one of ordinary skill in the art before the effective filing date.
Claim 12 is rejected under 35 U.S.C. 103 as being unpatentable over Church as evidenced by Kvitko et al. in view of Lin et al. and Harverd as applied to claims 1-3,6-9,11,17,18,20,21,31,34, 35,66-68 and 70-74 above, and further in view of Szybalski et al. (EP2423314, Published 29 Feb 2012).
The teachings of Church, Kvitko et al., Lin et al. and Harvard as applied to claims 1-3,6-9,11,17,18,20,21,31,34,35, 66-68 and 70-74 have been described above.
Church, Kvitko et al., Lin et al. and Harvard do not teach wherein the donor plasmid or recipient oligonucleotide comprises an inducible high-copy replication origin.
Before the effective filing date, Szybalski et al. taught the bacterial artificial chromosome (BAC) vectors were developed for the construction and faithful maintenance of genomic libraries in bacteria, and BAC vectors are based on the F plasmid, which maintains the vector at 1-2 copies per cell. Low copy number is an important feature for clone stability by limiting the amount of homologous sequence subject to recombination. However, the crucial disadvantage of low copy number is that it makes both preparing the vector for cloning and downstream analyses of clones, such as by sequencing and fingerprinting, more costly and laborious (paragraph 0003). Szybalski et al. taught medium- to high-copy number plasmids are often preferred over the low- or single-copy number plasmids for over-production of recombinant DNA and protein, because they typically lead to high yields of the target products (paragraph 0004). Szybalski et al. taught a vector may comprise an excisable fragment comprising an insertion site, a first and second origin of replication, and a pair of transcriptional terminators flanking the excisable fragment… the second origin of replication may be an inducible high-copy number origin of replication, such asoriV (paragraph 0007). Szybalski et al. taught the origin of replication may also be a high-copy origin of replication, which may not have a partitioning system. A high-copy origin of replication may segregate by mass action, which may cause empty cells to be minimized by the low probability in view of the number of vector copies. The high-copy origin of replication may maintain a high number of vectors in a host cell. The high-copy origin of replication may maintain 5-100 copies of the vector in a host cell. The high-copy origin of replication may also maintain more than 100 to more than 1000 copies of the vector in a host cell and a representative example of a high-copy origin of replication isoriV, and that a high-copy origin of replication may be used to increase production of the vector in a host cell.
It would have been obvious to one of ordinary skill in the art to have modified the donor plasmid used in the method of Church in view of Lin et al. and Harvard, to comprise an inducible high-copy replication origin based on the teachings of Szybalski et al. with a reasonable expectation of success. One of ordinary skill in the art would have been motivated to do so because Szybalski et al. taught the disadvantages of low copy number is that it makes both preparing the vector for cloning and downstream analyses of clones, such as by sequencing and fingerprinting, more costly and laborious, and that high-copy plasmids typically lead to high yields of the target products (paragraph 0004) and the high-copy origin of replication may maintain 5-100 copies or 100 to more than 1000 copies of the vector in a host cell and that a high-copy origin of replication may be used to increase production of the vector in a host cell.
Accordingly, the limitations of claim 12 would have been prima facie obvious to one of ordinary skill in the art.
Claim 13 is rejected under 35 U.S.C. 103 as being unpatentable over Church as evidenced by Kvitko et al. in view of Lin et al. and Harvard as applied to claims 1-3,6-9,11,17,18,20,21,31,34, 35,66-68 and 70-74 above, and further in view of Jones et al. (PLOS One July 2013, Vol. 8. Issue 7, e69319).
Claim Interpretation: The instant specification discloses the donor plasmid or recipient plasmid comprises a replicon that can replicate plasmids at least 30 kilobases in length (paragraph 0270). In further embodiments, the replicon is from a P1-derived artificial chromosome or a bacterial artificial chromosome (paragraph 0269). Therefore, art teaching a replicon from a P1-derived artificial chromosome or bacterial artificial chromosome reads on claim 13.
The teachings of Church, Kvitko et al., Lin et al. and Harvard as applied to claims 1-3,6-9,11,17,18,20,21,31,34,35,66-68 and 70-74 have been described above.
Church, Kvitko et al., Lin et al. and Harvard do not teach wherein the donor plasmid or recipient oligonucleotide comprises a replicon that can replicate plasmids of lengths greater than 30 kilobases.
Before the effective filing date, Jones et al. taught a procedure for the conjugative transfer of phage P1-derived Artificial Chromosome (PAC) library clones containing large natural product gene clusters (greater than or equal to 70 kilobases) to Streptomyces coelicolor strains that have been engineered for improved heterologous production of natural products (Abstract). Jones et al. taught while heterologous expression has been used successfully with small biosynthetic gene clusters (less than or equal to 40 kb), standard techniques are not as straightforward with large biosynthetic gene clusters (greater than or equal to 70 kb), largely because of the amount of DNA that needs to be cloned and transferred into a suitable expression host (introduction, left column). Jones et al. taught an approach which transfers large natural product gene clusters using phage P1-derived Artificial Chromosomes (PACs) for stable integration of PAC DNA via conjugation which creates a platform for gene cluster analysis, and enables targeted systems and synthetic biology approaches that may not be possible in wild-type strains.
It would have been obvious to one of ordinary skill in the art to have modified the donor plasmid used in the method of Church in view of Lin et al. and Harvard to comprise a P1-derived Artificial Chromosome (PAC) in order to replicate and transfer large natural product gene clusters based on the teachings of Jones et al. with a reasonable expectation of success, as this would amount to combining prior art elements according to known methods to yield predictable results. One of ordinary skill in the art would have been motivated to do so because Jones et al. taught that standard techniques are not as straightforward with large biosynthetic gene clusters (greater than or equal to 70 kb), because of the amount of DNA that needs to be cloned and transferred into a suitable expression host, and therefore taught an approach which transfers large natural product gene clusters using phage P1-derived Artificial Chromosomes (PACs) for stable integration of PAC DNA via conjugation which creates a platform for gene cluster analysis, and enables targeted systems and synthetic biology approaches that may not be possible in wild-type strains.
Accordingly, the limitations of claim 35 would have been prima facie obvious to one of ordinary skill in the art.
Claim 69 is rejected under 35 U.S.C. 103 as being unpatentable over Church as evidenced by Kvitko et al., in view of Lin et al. and Harvard as applied to claims 1-3,6-9,11,17,18,20,21,31,34, 35,66-68 and 70-74 above, and further in view of Menzel (US 20020102734, Published 01 Aug 2002).
The teachings of Church, Kvitko et al., Lin et al. and Harvard as applied to claims 1-3,6-9,11,17,18,20,21,31,34,35, 66-68 and 70-74 have been described above.
Church, Kvitko et al., Lin et al. and Harvard do not teach wherein each first donor cell is in a position in a first ordered array and each second, third or subsequent donor cell is in a position in a second, third or subsequent ordered array.
Before the effective filing date, Menzel taught methods and compositions for use of homologous recombination for directed evolution, gene reassembly, and directed mutagenesis and methods and compositions for use of bacterial conjugative transfer (paragraph 0002). Menzel taught conjugational gene transfer, a preferred procedure for transfer donor DNA into a target cell, is amenable to automation. Using liquid handling automation individual members of a donor library can be arrayed. Again employing liquid handling automation, an arrayed collection of donors may be individually mixed with a target (paragraph 0172). Menzel taught libraries suitable for the practice of directed gene assembly. Such libraries can be donor or vector libraries and can comprise a plurality of any of the donor or target vectors of the invention (paragraph 0175) and the vectors of the library are present within cells (e.g., donor cells for donor vectors and target cells for target vectors), e.g., bacterial cells (paragraph 0176). Menzel taught the members of the library are arrayed in a 96 (e.g., 8×12), 384 (e.g., 16×24), or a 1536 (e.g., 32×48) matrix or plate, e.g., microtiter plate or the donor library is present in a multiplicity of cells, e.g., bacterial cells, each cell containing a member of the library the members of which are arrayed and such cells are arrayed in a 96 (e.g., 8×12), 384 (e.g., 16×24), or a 1536 (e.g., 32×48) matrix or plate, e.g., microtiter plate (paragraph 0178).
It would have been obvious to one of ordinary skill in the art to modify the assembly method of Church in view of Lin et al. and Harvard such that the first donor cell is in a position in a first ordered array and each second, third or subsequent donor cell is in a position in a second third or subsequent donor array based on the teachings of Menzel with a reasonable expectation of success, as Menzel also pertains to homologous recombination and conjugative transfer in bacterial cells. One of ordinary skill in the art would have been motivated to do so because Menzel taught conjugational gene transfer, a preferred procedure for transfer donor DNA into a target cell, is amenable to automation and using liquid handling automation individual members of a donor library can be arrayed and an arrayed collection of donors may be individually mixed with a target (paragraph 0172). Menzel taught libraries suitable for the practice of directed gene assembly including donor or vector libraries and the vectors of the library are present within cells (e.g., donor cells for donor vectors and target cells for target vectors), e.g., bacterial cells (paragraph 0176) and the donor library is present in a multiplicity of cells, e.g., bacterial cells, each cell containing a member of the library the members of which are arrayed and such cells are arrayed in a 96 (e.g., 8×12), 384 (e.g., 16×24), or a 1536 (e.g., 32×48) matrix or plate, e.g., microtiter plate (paragraph 0178).
Accordingly, the limitations of claim 69 would have been prima facie obvious to one of ordinary skill in the art.
Claim 75 is rejected under 35 U.S.C. 103 as being unpatentable over Church as evidenced by Kvitko et al., in view of Lin et al. and Harvard as applied to claims 1-3,6-9,11,17,18,20,21,31,34, 35,66-68 and 70-74 above, and further in view of Dean et al. (US 20210324378, EFD 31 Oct 2018).
The teachings of Church, Kvitko et al., Lin et al. and Harvard as applied to claims 1-3,6-9,11,17,18,20,21,31,34,35, 66-68 and 70-74 have been described above.
Church, Kvitko et al., Lin et al. and Harvard do not teach wherein the method is used to construct combinatorial gRNA libraries.
Dean et al. taught compositions and methods for joining single-stranded and/or double stranded nucleic acid molecules permitting in vitro or in vivo assembly of multiple nucleic acid molecules with overlapping terminal sequences in a single reaction and methods and compositions can be useful for deterministic assembly of fragments of nucleic acid sequences (paragraph 0002). Dean et al. taught current pooled approaches for generating libraries are time-consuming and labor intensive, and expensive, and there is in a need in the art for new methods for generating complex nucleic acid assemblies, which do not suffer from the aforementioned drawbacks inherent with traditional methods for generating nucleic acid assemblies (paragraph 0005). Dean et al taught an assembly comprising a pair of first and second polynucleotides with an insert polynucleotide located therebetween generated using the methods and compositions provided herein is a guide RNA (gRNA) and in some cases, the methods provided herein are used to generate a library of gRNAs (paragraph 0140).
It would have been obvious to one of ordinary skill in the art before the effective filing date, to use the DNA assembly method of Church as evidenced by Kvitko et al. in view of Lin et al. and Harvard, to construct a combinatorial gRNA library based on the teachings of Dean et al. There would be a reasonable expectation of success as Dean et al. pertains to in vivo assembly of nucleic acid molecules. One of ordinary skill in the art would be motivated to do so because Dean et al. taught the need for new methods for generating complex nucleic acid assemblies that do not suffer from the drawbacks of current and traditional nucleic acid assembly methods, and that the methods and compositions can be used to generate a library of gRNAs.
Accordingly, the limitations of claim 75 would have been prima facie obvious to one of ordinary skill in the art.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to STEPHANIE L SULLIVAN whose telephone number is (703)756-4671. The examiner can normally be reached 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.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Ram R Shukla can be reached at 571-272-0735. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/STEPHANIE L SULLIVAN/Examiner, Art Unit 1635
/ABIGAIL VANHORN/Primary Examiner, Art Unit 1636
Prosecution Insights