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 .
Application Status
In response to Non-Final Office Action mailed on 05/26/2026, applicants' response, arguments and amendments filed on dated 08/26/2026 is acknowledged; in said response applicants’ have amended claims 34-35. Thus, amended claims 22-41 are pending in this application, Group II, claims 34-35 reading on the elected invention is now under consideration for examination; non-elected claims 22-33 and 36-41 of Groups' I and IIII are withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected invention, there being no allowable generic or linking claim. Applicant timely traversed the restriction (election) requirement in the reply filed on 05/08/2026. Rejections and/or objections not reiterated from previous office action are hereby withdrawn.
Withdrawn-Claim Rejections: 35 USC § 112(a)
Previous rejection of claims 34-35 rejected under 35 U.S.C. 112(a) for written-description and enablement, is being withdrawn due to claim amendments.
Withdrawn-Claim Rejections: 35 USC § 103
Previous rejection of claims 34-35 rejected under 35 U.S.C. 103(a) as being unpatentable over Mori et al., (US 12,241,102 B2; PCT Publication date 04/23/2020, filing date 10/18/2019) and further in view of Polturak et al., (PNAS., 2017, Vol. 114(34): 9062-9067), Wang et al., (Phytochem., 2016, Vol. 132: 16-25) and Larroude et al., (Microorganisms, 2021, Vol. 9, 838, pages 1-11), is being withdrawn due to claim amendments.
New-Claim Rejections: 35 USC § 103
Necessitated by claim amendments
The following is a quotation of 35 U.S.C. 103(a) which forms the basis for all obviousness rejections set forth in this Office action:
(a) A patent may not be obtained though the invention is not identically disclosed or described as set forth in section 102 of this title, if the differences between the subject matter sought to be patented and the prior art are such that the subject matter as a whole would have been obvious at the time the invention was made to a person having ordinary skill in the art to which said subject matter pertains. Patentability shall not be negatived by the manner in which the invention was made.
This application currently names joint inventors. In considering patentability of the claims under 35 U.S.C. 103(a), the examiner presumes that the subject matter of the various claims was commonly owned at the time any inventions covered therein were made absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and invention dates of each claim that was not commonly owned at the time a later invention was made in order for the examiner to consider the applicability of 35 U.S.C. 103(c) and potential 35 U.S.C. 102(e), (f) or (g) prior art under 35 U.S.C. 103(a).
The factual inquiries set forth in Graham v. John Deere Co., 383 U.S. 1, 148 USPQ 459 (1966), that are applied for establishing a background for determining obviousness under 35 U.S.C. 103(a) 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.
Claims 34-35 are rejected under 35 U.S.C. 103(a) as being unpatentable over Mori et al., (US 12,241,102 B2; PCT Publication date 04/23/2020, filing date 10/18/2019) and further in view of Polturak et al., (PNAS., 2017, Vol. 114(34): 9062-9067), Isayenkova et al., (Phytochem., 2006, Vol. 67: 1598-1612), Wang et al., (Phytochem., 2016, Vol. 132: 16-25) and Larroude et al., (Microorganisms, 2021, Vol. 9, 838, pages 1-11).
Regarding claims 34-35, Mori et al., (US 12,241,102 B2; PCT Publication date 04/23/2020, filing date 10/18/2019) teach yeast cell capable of producing one or more betalains, said yeast cell expressing: a first heterologous enzyme (TYH) capable of hydroxylating L-tyrosine and oxidizing L-DOPA; b. a second heterologous enzyme which is a 4,5-DOPA extradiol dioxygenase (DOD); and C. a third heterologous enzyme having glycosyltransferase activity; whereby said cell is capable of producing one or more betalains, wherein said one or more betalains comprise one or more glycosylated betalains. Mori et al., disclose a synthesis method for a betalain pigment, including: culturing a host that has been introduced therein the following gene encoding an amaranthin or gomphrenin-I-glucuronide synthetase, a gene encoding an enzyme having activity of hydroxylating a 3-position of a phenol ring of tyrosine, a gene encoding an enzyme having L-DOPA oxidase activity, a gene encoding an enzyme having activity of glycosylating a phenolic hydroxy group/glycosyltransferase, and a gene encoding an enzyme having DOPA 4,5-dioxygenase activity, and that has an ability to produce tyrosine or 3-hydroxy-L-tyrosine (L-DOPA), and extracting a betalain pigment from the host after the culturing (Abstract Fig. 1-3, reproduced below; and entire document);
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Mori et al., teach “The host to be used in the synthesis method for a betalain pigment of the present invention is not particularly limited, as long as the host has an ability to produce tyrosine or 3-hydroxy-L-tyrosine. For example, there may be used a synthesis system known per se, such as a recombinant Escherichia coli protein synthesis system, an insect protein synthesis system, a yeast protein synthesis system, a plant cell protein synthesis system, a cell-free protein synthesis system, a plant protein synthesis system, or animal cultured cells” (Col. 18, lines 28-38).
However, Mori et al., (US 12,241,102 B2; PCT Publication date 04/23/2020, filing date 10/18/2019) is silent regarding c. a third heterologous enzyme having glycosyltransferase activity, wherein said third heterologous enzyme is selected from: 1. Chenopodium quinoa glycosyltransferase CqSGT2 set forth in SEQ ID NO: 65, or a functional variant thereof having at least 80% sequence identity thereto; 2. Beta vulgaris glycosyltransferase BvSGT2 set forth in SEQ ID NO: 53, or a functional variant thereof having at least 80% identity thereto; 3. Bougainvillea glabra glycosyltransferase BgGT2 set forth in SEQ ID NO: 67, or a functional variant thereof having at least 80% identity thereto; and 4. Beta vulgaris glycosyltransferase BvSGT4 set forth in SEQ ID NO: 57, or a functional variant thereof having at least 80% identity thereto; and wherein said yeast cell comprises a mutation resulting in reduced activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD; as in claim 34).
Regarding claims 34-35, Polturak et al., (PNAS., 2017, Vol. 114(34): 9062-9067) also teach a method for producing one or more betalains/biosynthetic pathway, expression of three genes comprising a first heterologous enzyme (TYH) capable of hydroxylating L-tyrosine and oxidizing L-DOPA; b. a second heterologous enzyme which is a 4,5-DOPA extradiol dioxygenase (DOD); and C. a third heterologous enzyme having glycosyltransferase activity (Abstract; col. 2, ¶ 2, page 9062; Fig. 1, page 9063; Fig. 4A, page 9065; and entire document).
Regarding claim 34, i.e., c. a third heterologous enzyme having glycosyltransferase activity, wherein said third heterologous enzyme is selected from: 1. Chenopodium quinoa glycosyltransferase CqSGT2 set forth in SEQ ID NO: 65, or a functional variant thereof having at least 80% sequence identity thereto; 2. Beta vulgaris glycosyltransferase BvSGT2 set forth in SEQ ID NO: 53, or a functional variant thereof having at least 80% identity thereto; 3. Bougainvillea glabra glycosyltransferase BgGT2 set forth in SEQ ID NO: 67, or a functional variant thereof having at least 80% identity thereto; and 4. Beta vulgaris glycosyltransferase BvSGT4 set forth in SEQ ID NO: 57, or a functional variant thereof having at least 80% identity thereto; Isayenkova et al., (Phytochem., 2006, Vol. 67: 1598-1612), provide the structural and functional elements of the instant invention. Isayenkova et al., disclose regio-selective flavonoid UDP-glucosyltransferases having 99.3% sequence identity to SEQ ID NO: 53 and 81.3% sequence identity to SEQ ID NO: 65 obtained from Beta vulgaris of the instant invention; said reference also provides evidence said reference glucosyltransferases having the activity of glycosylation of betanidin (see Abstract; Table 2 & Fig. 3, page 1601; Fig. 4, page 1602; Table 3, page 1603; and entire document).
Regarding claim 34, i.e., yeast cell comprises a mutation resulting in reduced activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD) the following references via different lines of evidence provide teaching, suggestion and motivation to reduce/eliminate the activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD) in a host cell engineered to produce betalains for the following reasons:
Wang et al., (Phytochem., 2016, Vol. 132: 16-25) provide teaching, suggestion and motivation to reduce/eliminate the activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD), as 4-HPPD is involved in directing the carbon flow 4-hydroxyphenylpyurvate (4-HPP) towards unwanted by-product/pigment homogentisate (see Abstract; Fig. 1, page 17); and 4-hydroxyphenylpyurvate (4-HPP) is an intermediate necessary for the production of betalain (Fig. 1, page 17), and therefore a skilled artisan would be motivated to reduce the activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD) to increase the production of betalain.
Larroude et al., (Microorganisms, 2021, Vol. 9, 838, pages 1-11) also provide teaching, suggestion and motivation to reduce/eliminate the activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD), as 4-HPPD is involved in directing the carbon flow 4-hydroxyphenylpyurvate (4-HPP) towards unwanted by-product/pigment homogentisate (see Abstract; Fig. 1, page 2) and provide evidence in a yeast cell/ Yarrowia lipolytica deletion of hydroxyphenylpyruvate dioxygenase (4-HPPD) resulted in elimination of by-product/pigment homogentisate (Table 1, DJMY8131 strain Yarrowia lipolytica deletion of hydroxyphenylpyruvate dioxygenase (4-HPPD), Fig. 4, page 11).
Hence, a skilled artisan will be motivated to reduce the activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD), said metabolic engineering in yeast involves the modification of endogenous biosynthetic pathways to increase the biosynthesis of target bioactive compounds/betalain, reduce the production of an unwanted compound/homogentisate or other metabolites and unwanted pigments. A skilled artisan will be motivated in a single step modification of a biosynthetic pathway or competitive pathways blocked to divert metabolic flow towards betalain formation. The identification of the genes encoding enzymes involved in the betalain biosynthetic pathway by Mori et al., and Polturak et al., will motivate the skilled artisan for the development of metabolic engineering strategies to increase betalain production by reducing/eliminating the activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD) as described in references of Wang et al., and Larroude et al.
As such, disclosure of strategy and methods including structural and functional elements for a method of producing betalain in the claimed yeast and process, such as that of references of Mori et al., Polturak et al., Isayenkova et al., Wang et al., and Larroude et al., teaching the advantages of said modifications, and clearly suggests to a skilled artisan to modify the teachings of Mori et al., and Polturak et al., and a skilled artisan would be motivated to incorporate the structural and functional elements of the cited secondary references of Isayenkova et al., Wang et al., and Larroude et al., in the claimed yeast and process, and the combined references also provide the advantages of modifying the primary teachings of Mori et al., by incorporating the structural and functional elements of secondary references as claimed in the instant invention and is well known in the art (for details see the rejection above).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claimed process, as suggested by Mori et al., Polturak et al., Isayenkova et al., Wang et al., and Larroude et al. A person of ordinary skill in the art is motivated to make such change depending on the experimental need i.e., yeast cell capable of producing one or more betalains, said yeast cell expressing: a. a first heterologous enzyme (TYH) capable of hydroxylating L-tyrosine and oxidizing L-DOPA; b. a second heterologous enzyme which is a 4,5-DOPA extradiol dioxygenase (DOD); and C. a third heterologous enzyme having glycosyltransferase activity; whereby said cell is capable of producing one or more betalains, wherein said one or more betalains comprise one or more glycosylated betalains, and wherein said yeast cell comprises a mutation resulting in reduced activity of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD) (as in claims 34-35), because a skilled artisan would realize such a modification would be useful to efficiently produce betalain in the claimed process. One of ordinary skill in the art has a reasonable expectation of success at adding the steps in the claimed yeast and process, as suggested in the teachings of Polturak et al., Wang et al., and Larroude et al., and are well known in the art. Therefore, the invention as a whole lack an inventive step over the prior art. The expectation of success is high, because the combined teachings of Mori et al., Polturak et al., Isayenkova et al., Wang et al., and Larroude et al., also provide the structural and functional elements of the instant invention (Teaching, Suggestion and Motivation).
Given this extensive teaching in prior art (Mori et al., Polturak et al., Isayenkova et al., Wang et al., and Larroude et al.,) the claimed yeast and process, as taught by the instant invention and as claimed in claims 34-35 is not of innovation but of ordinary skill in the art and the expectation of success is extremely high i.e., “a person of ordinary skill has good reason to pursue the known options within his or her technical grasp. If this leads to the anticipated success, it is likely that product [was] not of innovation but of ordinary skill and common sense. In that instance the fact that a combination was obvious to try might show that it was obvious under § 103.”KSR, 550 U.S. at, 82 USPQ2d at 1397”.
Hence, claims 34-35 are rejected under 35 U.S.C. 103(a) as being unpatentable over Mori et al., (US 12,241,102 B2; PCT Publication date 04/23/2020, filing date 10/18/2019) and further in view of Polturak et al., (PNAS., 2017, Vol. 114(34): 9062-9067), Isayenkova et al., (Phytochem., 2006, Vol. 67: 1598-1612), Wang et al., (Phytochem., 2016, Vol. 132: 16-25) and Larroude et al., (Microorganisms, 2021, Vol. 9, 838, pages 1-11).
Applicants’ have traversed the above 35 USC § 103 following claim amendments with the following arguments (see pages 22-24 of Applicants’ REMARKS dated 08/26/2026).
Applicants’ argue: “…Accordingly, a prima facie case of obviousness has not been established because the cited reference combination does not teach or suggest the claimed heterologous enzymes having glycosyltransferase activity expressed by a yeast cell capable of producing one or more betalains. The cited references do not provide one of ordinary skill in the art a reasonable expectation of success to substitute the claimed scopoletin glucosyltransferase enzyme sequences to arrive at the claimed yeast cell…”.
Reply: Applicants' arguments have been considered but are found to be non-persuasive for the following reasons. Examiner continues to maintain the rejection for reasons stated on record (dated 05/26/2026) and additionally for the following reasons. Contrary to applicants’ arguments, examiner has provided a new reference Isayenkova et al., (Phytochem., 2006, Vol. 67: 1598-1612), provide the structural and functional elements of the instant invention. Isayenkova et al., disclose regio-selective flavonoid UDP-glucosyltransferases having 99.3% sequence identity to SEQ ID NO: 53 and 81.3% sequence identity to SEQ ID NO: 65 obtained from Beta vulgaris of the instant invention; said reference also provides evidence said reference glucosyltransferases having the activity of glycosylation of betanidin (see Abstract; Table 2 & Fig. 3, page 1601; Fig. 4, page 1602; Table 3, page 1603; and entire document.
Contrary to applicants’ arguments, examiner continues to maintain that Mori et al., (US 12,241,102 B2; PCT Publication date 04/23/2020, filing date 10/18/2019) teach yeast cell capable of producing one or more betalains, said yeast cell expressing: a first heterologous enzyme (TYH) capable of hydroxylating L-tyrosine and oxidizing L-DOPA; b. a second heterologous enzyme which is a 4,5-DOPA extradiol dioxygenase (DOD); and C. a third heterologous enzyme having glycosyltransferase activity; whereby said cell is capable of producing one or more betalains, wherein said one or more betalains comprise one or more glycosylated betalains: (see rejection above) and additionally applicants’ are also directed to Fig. 16 in Mori et al., (US 12,241,102 B2) reproduced below:
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Furthermore, it is well known in the art that Betalains (also called betacyanins) include the reddish to violet betalain pigments. Among the betalains present in plants include betanin, isobetanin, probetanin, and neobetanin (see enclosed evidentiary reference Wikipedia-Betalain, 6 pages downloaded on 09/05/2026).
Summary of Pending Issues
The following is a summary of issues pending in the instant application.
Claims 34-35 are rejected under 35 U.S.C. 103(a) as being unpatentable over Mori et al., (US 12,241,102 B2; PCT Publication date 04/23/2020, filing date 10/18/2019) and further in view of Polturak et al., (PNAS., 2017, Vol. 114(34): 9062-9067), Isayenkova et al., (Phytochem., 2006, Vol. 67: 1598-1612), Wang et al., (Phytochem., 2016, Vol. 132: 16-25) and Larroude et al., (Microorganisms, 2021, Vol. 9, 838, pages 1-11).
Claims 22-33 and 36-41 are withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected invention.
Conclusion
None of the claims are allowable. Claims 34-35 are rejected for the reasons identified in the Rejections and Summary sections of this Office Action. Applicants must respond to the rejections in each of the sections in this Office Action to be fully responsive for prosecution.
THIS ACTION IS MADE FINAL. Applicants are 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 extension fee 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.
Regarding filing an After Final amendment, Applicants are directed to MPEP 714.13, which states:
II. ENTRY NOT A MATTER OF RIGHT
It should be kept in mind that applicant cannot, as a matter of right, amend any finally rejected claims, add new claims after a final rejection (see 37 CFR 1.116) or reinstate previously canceled claims. Except where an amendment merely cancels claims, adopts examiner suggestions, removes issues for appeal, or in some other way requires ONLY A CURSORY REVIEW by the examiner (e.g., typographical errors), compliance with the requirement of a showing under 37 CFR 1.116(b)(3) is expected in all amendments after final rejection. An affidavit or other evidence filed after a final rejection, but before or on the same date of filing an appeal, may be entered upon a showing of good and sufficient reasons why the affidavit or other evidence is necessary and was not earlier presented in compliance with 37 CFR 1.116(e). See 37 CFR 41.33 and MPEP § 1206 for information on affidavit or other evidence filed after appeal. (Examiner's emphasis) If more than a cursory review is required, Applicants are referred to CFR §1.114.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to GANAPATHIRAMA RAGHU whose telephone number is (571)272-4533. The examiner can normally be reached on M-F 8:30am-5pm EST.
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/GANAPATHIRAMA RAGHU/ Primary Examiner, Art Unit 1652
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