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
The Amendments and Remarks filed 06 July 2026 in response to the Office Action 07 January 2026 are acknowledged and have been entered. Claims 1 and 11 are amended. Claims 1-19 are pending and being examined on the merits.
Any objection or rejection not reiterate herein has been overcome by applicants claim amendments.
Priority
This application is a divisional of application 16/533,928 filed 08/07/2019 which claims priority to application 62/715,498 filed 08/07/2018.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1, 3-8, 10-16 and 19 are rejected under 35 U.S.C. 103 as being unpatentable over Thorsness (US 2014/0302577 A1) in view of Sharma (Sharma and Liebman. Molecular Microbiology (2012) 86(4), 866–881) and Magalhães (Magalhães et. al. 2017. J Ind. Microbiol. Biotechnol. 44:1203–1213). This is a new rejection necessitated by applicants claim amendments.
Regarding claims 1, 3-4 and 11, Thorsness teaches methods for enhancing yeast fermentation of plant material through the genetic modifications of non-respiring yeast, i.e. Saccharomyces cerevisiae, through the alteration of yeast nuclear or mitochondrial genes required for growth on non-fermentable carbon sources [0042]. Thorsness teaches that transgenic yeast of the present disclosure having a nonfunctional or absent mitochondrial DNA express enhanced fermentation and improved growth [0042]. Thorsness teaches that yeast strains with intact, fully-functional mtDNA (ρ+ strains) can be converted into strains without mtDNA (ρ° strains) or with dysfunctional mtDNA (ρ- strains) by inclusion of ethidium bromide (EtBr), i.e. a mitochondrial elimination agent, in the growth media [0046]. Thorsness teaches the mitochondrial DNA of an industrial strain diploid yeast is removed from the diploid yeast by growing the diploid yeast in the presence of ethidium bromide [0048, Fig. 2]. Thorsness teaches crossing/mating a kar1-1 strain bearing a mitochondria genome (i.e., second yeast strain) with the diploid yeast strain lacking mitochondria (i.e., first yeast strain) where the mitochondria from the kar1-1 strain was introduced into the yeast strain lacking mitochondria [0048; Fig. 2]. Thorsness teaches the selection of the yeast that contains the nuclear genome of the diploid strain (i.e., first yeast strain) and mitochondria genome of the kar1-1 strain (i.e., second yeast strain) and where 100% of the yeast are from the second yeast strain (regarding claim 11) [0048]. Thereby, Thorsness teaches methods for producing genetically modified yeast strains by replacing mitochondrial genomes through cytoduction using kar1-mediated mitochondrial transfer after generating mitochondrial DNA-deficient recipient strains. Thorsness explains that mitochondrial DNA can be removed from a recipient yeast strain to generate cells lacking mtDNA (rho0 cells), followed by introduction of donor mitochondrial genomes into those recipient strains by cytoduction using kar1 mutants to inhibit nuclear fusion, thereby producing strains possessing the recipient nuclear genome and donor mitochondrial genome.
Regarding claim 5, the teaching of the mitochondria coming from the kar1-1 strain to the diploid strain is a teaching where the mitochondria are from a donor yeast strain [0048].
Regarding claims 6 and 10, Thorsness teaches that the yeast strains are the Saccharomyces cerevisiae strain.
Thorsness does not explicity teach that the donor strain comprises functional mitochondria, nor selecting for functional mitochondria after mating. Thorsness does not teach where the first yeast strain is a Saccharomyces cerevisiae x Saccharomyces eubayanus hybrid, where the first yeast strain is a lager-brewing strain or where at least 90% of the mitochondria are from Saccharomyces cerevisiae or Saccharomyces eubayanus.
Sharma teaches cytoduction that is carried out with RHO+ donors and mitochondria deficient (rho-) recipients [pg. 879, col. 1, para 3]. Sharma teaches that either donor or recipient carried a kar1 mutation that inhibits nuclear fusion [pg. 879, col. 1, para 3]. Sharma teaches that following mating, cytoductants and diploids were selected by growth on synthetic media lacking amino acids specifically required by the donor and using glycerol as the sole carbon source where functional mitochondria are required for growth; thus, the cytoductant would inherit the nucleus from one parent and mitochondria from another [pg. 879, col. 1, para 3]. Thereby, Sharma teaches the production of a cybrid yeast strain comprising the genotype of the first yeast strain and the mitotype of the second yeast strain where the second yeast strain comprises a function mitochondria and selecting the yeast for functional mitochondria.
Magalhães teaches a hybrid yeast generated by crossing the cryotolerant Saccharomyces eubayanus with a Saccharomyces cerevisiae wine strain and assessed the suitability of the hybrids for low-temperature cider fermentation [abstract]. Magalhães teaches that the hybrid strains outperformed Saccharomyces cerevisiae , which was sensitive to low temperatures [abstract]. Magalhães teaches that 37°C is a restrictive temperature for Saccharomyces eubayanus and they are unable to grow [pg. 1204, col. 2, para 2; Fig. 1]. Magalhães teaches that mitochondria inheritance can influence the phenotypic properties (i.e. temperature tolerance) of new hybrids [pg. 1210, col. 2, para 2]. Magalhães further teaches where the temperature employed in alcoholic beverages fermentations impacts the sensorial properties of the final product and that there are several reasons why cold fermentations (i.e., lower than 15 °C) are preferred over warm fermentations and that it is necessary to have yeast strain that is able to survive and retain metabolic activity at these temperatures if fermentation is to proceed efficiently [pg. 1203, col. 2, para 1]. Magalhães teaches that interspecific hybridization can apparently be used effectively to improve low-temperature fermentation performance (i.e. increased cold tolerance) without compromising product quality [abstract]. Magalhães teaches the lager yeast S. cerevisiae × S. eubayanus hybrids [pg. 1203, col. 2, para 2] (regrading claim 8). Magalhães additionally teaches that naturally cold-tolerant strains like Saccharomyces eubayanus tend to have higher ethanol sensitivity (a phenotypic property) than Saccharomyces cerevisiae and may, therefore, be less suitable for alcoholic fermentation [pg. 1203, col. 2, para 2]. Magalhães teaches that Saccharomyces cerevisiae was a wine strain and that cider is lower in alcohol than wine [pg. 1204, col.2, para 1-2]. Magalhães also teaches that the hybrid C967 plotted closely to the S. cerevisiae parent, and produces a compound with higher alcohols [Fig. 4]. Thereby, Magalhães that hybrids inherit advantageous mitochondria characteristics resulting in superior fermentation phenotypes relative to parental strains. Magalhães further teaches teaches evaluating mitochondrial origin and confirms mitochondrial functionality by plating strains on glycerol medium, where respiration-competent mitochondria are required for growth [pg. 1205, col. 2, para 2]. Accordingly, Magalhães teaches that mitochondrial genotype is an important determinant of industrially useful fermentation traits and that respiratory growth on glycerol is an accepted method of confirming mitochondrial functionality.
Regarding claim 14 and 16, Magalhães teaches a method of fermenting juice to cider (i.e., an alcoholic beverage) using the hybrid yeast strains at different temperatures [pg. 1207 last paragraph – pg. 1208, col. 1, para 1; Fig. 3].
Regarding claim 19, Magalhães teaches a method of fermenting juice at temperatures below 60°F [pg. 1206, col. 1, para 1-2; Fig. 3].
Regarding claim 1 and 11, it would have been obvious to one ordinary skilled in the art before the effective filing date of the claimed invention to modify the mitochondrial transfer method of Thorsness by employing the conventional cytoductant selection procedure taught by Sharma and applying the resulting cybrid production methods to obtain yeast strains having advantageous mitochondrial phenotypes as taught by Magalhães. Sharma teaches that selection on glycerol medium is a standard technique for recovering only those cytoductants that have successfully inherited functional donor mitochondria, thereby eliminating mitochondrial-deficient recipients. Magalhães teaches that mitochondrial inheritance significantly influences desirable fermentation characteristics such as cryotolerance and fermentation efficiency and that mitochondrial functionality is routinely evaluated by growth on glycerol medium. Therefore, one of ordinary skill therefore would have been motivated to employ Sharma’s established cytoductant-selection procedure in the mitochondrial replacement system of Thorsness in order to efficiently recover cybrid yeast strains possessing functional donor mitochondria suitable for the improved fermentation characteristics sought by Magalhães, with a reasonable expectation of success because each reference uses conventional Saccharomyces cytoduction methodologies.
Regarding claims 7-8, it would have been obvious to one ordinary skilled in the art before the effective filing date of the claimed invention to modify the method of Thorsness where the first yeast strain is a Saccharomyces cerevisiae x Saccharomyces eubayanus hybrid. One of ordinary skill would be motivated to make the modification for the advantage of generating a yeast strain containing Saccharomyces cerevisiae mitochondria that are capable of fermentations at higher temperatures and for producing wine and drinks with higher alcohol concentrations than cider.
Regarding claims 12-13, it would have been obvious to one ordinary skilled in the art before the effective filing date of the claimed invention to modify the method of Thorsness by treating either the Saccharomyces cerevisiae strain or the Saccharomyces eubayanus strain with EtBr to produce a corresponding mitochondria-null yeast strain prior to mating that strain with a kar1-1 deficient Saccharomyces eubayanus strain or Saccharomyces cerevisiae strain, respectively, and select for yeast that contains the nuclear genome of the first yeast strain and mitochondria genome of the kar1-1 strain (i.e., at least 90% of the mitochondria). One of ordinary skill would be motivated to make the modification for the advantage of assessing each hybrid’s ability to enhance yeast fermentation. The combination of prior art elements according to known methods to yield predictable results supports can support a conclusion of obviousness. See MPEP 2143(I). One of ordinary skill in the art would have a reasonable expectation of success since Thorsness, Magalhães, and Sharma each teach the production of yeast strains with altered mitotypes for growth and fermentation advantages.
Claim 2 is rejected under 35 U.S.C. 103 as being unpatentable over Thorsness (US 2014/0302577 A1) in view of Sharma (Sharma and Liebman. Molecular Microbiology (2012) 86(4), 866–881) and Magalhães (Magalhães et. al. 2017. J Ind. Microbiol. Biotechnol. 44:1203–1213) as applied to claim 1 and further in view of Alexander (US 2018/0127784 A1).
The teachings of Thorsness, Magalhães and Sharma are discussed above as applied to claim 1 and similarly apply to claim 2.
Thorsness, Magalhães and Sharma do not teach wherein the first mitochondria-null yeast strain comprises a HyPr plasmid.
Alexander teaches a generalized method for the efficient production of designer hybrid strains of Saccharomyces based on a series of inducible expression plasmids named HyPr (for Hybrid Production) [0083]. Alexander teaches that these plasmids contain complementary dominant drug-resistance markers, a doxycycline-inducible HO cassette, and a generalized replication origin that provides functionality across Saccharomyces and many other yeasts [0083]. Alexander teaches that HyPr efficiently produces allotetraploid and autotetraploid strains of Saccharomyces, as well as allohexaploid strains and strains of higher ploidies [0083]. The resulting strains can be rapidly screened for plasmid loss [0083].
It would have also been obvious to one ordinary skilled in the art before the effective filing date of the claimed invention to modify the method as taught and suggested by Thorsness where the first mitochondria-null yeast strain comprises a HyPr plasmid. One of ordinary skill would be motivated to make the modification for the advantage of screening or selection of the strains comprising the altered mitotype.
Claims 9, 15 and 17 are rejected under 35 U.S.C. 103 as being unpatentable over Thorsness (US 2014/0302577 A1) in view of Sharma (Sharma and Liebman. Molecular Microbiology (2012) 86(4), 866–881) and Magalhães (Magalhães et. al. 2017. J Ind. Microbiol. Biotechnol. 44:1203–1213) as applied to claim 1, 7, 8, and 14 and further in view of Nakao (WO2007/099451).
The teachings of Thorsness, Magalhães and Sharma are discussed above as applied to claim 1, 7, 8, and 14 and similarly apply to claim 9, 15, and 17.
Thorsness, Magalhães and Sharma do not teach the first yeast strain is Saccharomyces pastorianus or Saccharomyces carlsbergensis. Magalhães teaches the lager yeast S. cerevisiae × S. eubayanus hybrids [pg. 1203, col. 2, para 2]; however, Thorsness, Magalhães and Sharma do not teach wort as a fermentable substrate or where the fermentation product is lager.
Nakao teaches that specific examples of yeast available for fermentation, for example brewery yeast for beer, wine and sake, include yeast of genus Saccharomyces, brewery yeast such as Saccharomyces pastorianus, Saccharomyces carlsbergensis, and Saccharomyces cerevisiae [pg. 27, last paragraph]. Nakao teaches that lager beer is fermented using bottom fermenting yeast belonging to Saccharomyces pastorianus using wort extract [pg. 10, lines 23-25; example 1].
It would have also been obvious to one ordinary skilled in the art before the effective filing date of the claimed invention to modify the method as taught and suggested by Thorsness and Magalhães to where the first yeast strain is Saccharomyces pastorianus for the production of lager beer using wort extract. One of ordinary would be motivated to make the modification given the teachings of Nakao who teaches that lager beer is fermented using bottom fermenting yeast belonging to Saccharomyces pastorianus using wort extract and Magalhães who teach that S. cerevisiae × S. eubayanus hybrids are lager yeast. This modification would amount to the substitution of one known lager yeast for another.
Response to Arguments
Applicant’s arguments, see pages 6-8, filed 06 July 2026, with respect to the rejection(s) of claims 1, 3-6 and 10-11 under 35 U.S.C. 102(a)(1), and claims 1-17 and 19 under 35 U.S.C. 103 have been fully considered and are persuasive. Therefore, the rejections have been withdrawn. However, upon further consideration, a new ground(s) of rejection is made in view of Sharma. The combined teachings of Thorsness, Sharma, and Magalhaes collectively teach each limitation of the amended claims and would have suggested their combination to one of ordinary skill in the art for the predictable purpose of producing cybrid yeast strains possessing functional donor mitochondria and desirable fermentation characteristics.
Allowable Subject Matter
The following is a statement of reasons for the indication of allowable subject matter: Claim 18 is free of the art because the prior art does not teach or suggest fermentation at a temperature at or above 60°F for the claimed cybrids.
Claim 18 is objected to as being dependent upon a rejected base claim, but would be allowable if rewritten in independent form including all of the limitations of the base claim and any intervening claims.
Conclusion
No claims 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 TIFFANY N GROOMS whose telephone number is (571)272-3771. The examiner can normally be reached on M-F 830-530.
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/T.N.G./Examiner, Art Unit 1637 /Jennifer Dunston/Supervisory Patent Examiner, Art Unit 1637
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