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 .
Claims 2-4, 13, 14 have been canceled. Claims 1, 5-12, 15, 17-19 remain pending and under consideration.
Applicant's arguments filed 1-30-26 have been fully considered but they are not persuasive.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Claim Objections
Claim 1 is missing the fact that the fish is genetically modified.
If the inventive concept is that albinism in [red] tilapia was unexpected when inactivating an endogenous slc45a2 gene, then claim 1 can be written more simply and accurately. Assuming “albino” means the skin of the fish completely lacks pigment and the fish has red eyes, it would be clearer to say ---A genetically modified albino tilapia whose genome comprises a homozygous inactivated slc45a2 gene---.
Claim Rejections - 35 USC § 103
Claims 1, 5-12, 15, 17-19 remain rejected under 35 U.S.C. 103 as being unpatentable over Edvardson (PLoS, 2014, Vol. 9, No. 9, e108622, pg 1-7), Shiraki (Scientific Reports, 2018, Vol. 8, No. 13366, pg 1-15), Li (Genetics, 2014, Vol. 197, pg 591-599), Zhu (Scientific Reports, 2016, Vol. 6, No. 31347, pg 1-12), Bian (Marine Drugs, 2019, Vol. 17, No. 386, pg 1-14).
Edvardson taught a genetically modified founder salmon whose genome comprises an inactivated slc45a2 gene (abstract, materials and methods, discussion). The salmon had a “graded phenotype ranging from complete lack of pigmentation to partial loss and normal pigmentation” (abstract) with red eyes (pg 3, Fig. 1A). The founder salmon were bred to homozygosity (pg 2, line 2). The “complete lack of pigmentation” with red eyes or the “partial loss” of pigmentation and red eyes is equivalent to “a heritable albino-red phenotype characterized by a red eye and skin phenotype and complete absence of black/grey pigmentation in skin, peritoneum and eyes”. Edvardson and Shiraki bred the fish for numerous generations (see citations above) so the mutation is “heritable” as required in claim 4. Edvardson bred the fish for numerous generations (see citations above) so the mutation is “heritable” as required in claim 1.
Shiraki taught a genetically modified founder zebrafish whose genome comprises an inactivated slc45a2 gene (abstract, materials and methods, discussion). The zebrafish were albino (e.g. pg 9, “Fluorescence imaging in the head of Tg-albino”) with red eyes (pg 8, Fig. 6F, 6G). Shiraki founder zebrafish were bred to homozygosity (pg 9, last partial paragraph). The “complete lack of pigmentation” with red eyes is equivalent to “a heritable albino-red phenotype characterized by a red eye and skin phenotype and complete absence of black/grey pigmentation in skin, peritoneum and eyes”. Shiraki bred the fish for numerous generations (see citations above) so the mutation is “heritable” as required in claim 1.
The transgenic slc45a2 founder fish of Edvardson and Shiraki are “hybrid” fish as required in claim 1 because the cells of the fish have different genomes – the germ cells have the transgene in their genome, but not all somatic cells have the transgene in their genome. They do not have “said mutation [ ] in a homozygous form” as required in claim 1.
The homozygous slc45a2 transgenic fish of Edvardson and Shiraki are “hybrid” fish as required in claim 1 because they have endogenous DNA and exogenous DNA in their genome. They have “said mutation [ ] in a homozygous form” as required in claim 1 because ALL cells of the homozygous transgenic fish have the transgene in their genome.
Edvardson and Shiraki did not teach inactivating the slc45a2 gene in tilapia as required in claim 1.
However, the means for inactivating a variety of target genes by identifying gRNA target sequences, making Cas9 mRNA, and microinjecting gRNA and Cas9 mRNA into tilapia embryos, and obtaining tilapia with knocked out genes was well known as shown by Li (pg 592, “gRNA design…”, “Cas9 messenger mRNA…”, “Microinjection, genomic DNA extraction, and mutation detection assay”; pg 593, Table 1, nanos2, nanos3, dmrt1, foxl2). A transcriptome analysis of genes (including slc45a2) related to skin color in tilapia was described by Zhu (throughout; pg 6, halfway down; pg 7, 2nd full para). The genome sequence of tilapia was well-known as shown by Bian (“Whole genome sequencing of the blue tilapia…” - title).
Thus, it would have been obvious to those of ordinary skill in the art at the time of filing to inactivate an slc45a2 gene in a fish as described by Edvardson and Shiraki in tilapia as described by Li. Those of ordinary skill in the art at the time of filing would have been motivated to do so to understand how the slc45a2 gene affects the color pattern in tilapia.
Edvardson taught injecting 2-3 hour old fertilized eggs and allowed them to develop into a fish (pg 2, col. 2, “Injection procedures”). Shiraki taught injecting one-cell fertilized eggs (pg 12, “generation of transgenic albino (Tg-albino) fish”) and allowed them to develop. Li injected one-cell stage embryos (pg 592, col. 2, “Microinjection…”) and allowed them to develop into a fish. This is equivalent to “introducing into a zygote [ ] a DNA editing agent” into a zygote and allowing them to develop into a fish as required in claim 5.
Edvardson, Shiraki, and Li all taught how to identify whether the desired inactivation occurred (see citations above) which is equivalent to “identify[ing] said [tilapia] comprising the loss-of-function mutation” as required in claim 5.
Edvardson, Shiraki, and Li all bred a founder fish with the desired inactivation (see citations above) which is equivalent to “breeding the fish [ ] with a second fish [ ] to produce a third fish” as required in claim 5.
Edvardson, Shiraki, and Li inbred F1 fish with the desired inactivation (see citations above) which is equivalent to “the first fish and the second fish both carry at least one allele with a loss-of-function mutation” as required in claim 5.
Edvardson, Shiraki, and Li inbred F1 fish to obtain F2 fish with a homozygous disruption of the desired inactivation (see citations above) which is equivalent to “the third fish is homozygous for the loss-of-function mutation” as required in claim 5.
Edvardson, Shiraki, and Li all taught deletions (see citations above) as required in claim 10.
Edvardson, Shiraki, and Li all taught deletions (see citations above) that span a number of introns and exons which is equivalent to “two or more mutations” as required in claim 11.
Claim 12 has been included because while SEQ ID NO: 9-12 correspond to SEQ ID NO: 1, they are deleted from the genome of the fish, so they do not exist in the fish. If applicants are attempting to further limit the target sequences for gRNA, then much clarification is required. Claim 9 has also been included because SEQ ID NO: 9 was part of the genome of numerous fish:
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The starting fish described by Edvardson, Shiraki, and Li are all purebred as required in claim 15. The F2 homozygous fish described by Edvardson, Shiraki, and Li are also “purebred” because they are homozygous for the inactivated target gene.
Claim 17 has been included because Edvardson taught targeting two gRNA sequences:
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and Shiraki taught targeting two gRNA sequences:
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Edvardson, Shiraki, and Li all taught “progeny” of the genetically modified fish as required in claim 18.
Any of the fish described by Edvardson, Shiraki, and Li are “feed or food product” as required in claim 19.
Response to arguments
Applicants argue the motivational statement requires hindsight. Applicants’ argument is not persuasive. It must be recognized that any judgment on obviousness is in a sense necessarily a reconstruction based upon hindsight reasoning. But so long as it takes into account only knowledge which was within the level of ordinary skill at the time the claimed invention was made, and does not include knowledge gleaned only from the applicant's disclosure, such a reconstruction is proper. See In re McLaughlin, 443 F.2d 1392, 170 USPQ 209 (CCPA 1971). . The results will facilitate understanding the molecular mechanisms of skin pigmentation differentiation in red tilapia and accelerate the molecular selection of the specific strain with consistent skin colors.
Applicants argue the phenotype in albino was unexpected. Applicants’ argument is not persuasive because the exact phenotype claimed was obtained in two other types of fish by inactivating a slc45a2 gene. Applicants’ argument is also not persuasive because it is flawed. It fails to acknowledge what was expected as the basis of the argument, i.e. two types of transgenic fish with an inactivated slc45a2 gene had the phenotype claimed. It fails to account for any secondary considerations, i.e. transgenic tilapia were well-known. It fails to properly compare what was expected to applicants results. Therefore, the “unexpected results” argument is legally, logically, and scientifically flawed.
Applicants argue “Zhu et al. directly contradicts the claimed invention by teaching that slc45a2 contributes to BLACK coloration in red tilapia, specifically stating: "The expression level of tyrosinase-related protein 1 (tyrpl), sex determining region Y-box 10 (sox10), premelanosome protein (pmel/silv), solute carrier family 24 (sodium/potassium/calcium exchanger) member 5 (slc24a5), solute carrier family 45 member 2 (slc45a2) and tyrosinase (tyr) were up-regulated in PB skin, implying that these genes play a key role in the contribution to black coloration in red tilapia in the eumelanin synthesis pathway." Critically, only one of three red tilapia strains (PB - pink with scattered black spots) showed slc45a2 upregulation, while the other two red strains tested showed no such upregulation, demonstrating that slc45a2 is not consistently associated with red coloration in tilapia.” Applicants’ argument is not persuasive because Edvardson used salmon which are red and made them albino.
Applicants argue the references don’t teach lack of pigment in the tissues claimed. Applicants’ argument is not persuasive because the references say they observed complete lack of pigment or albinism.
Applicants cite Glasauer and Rohner and argue there is a phylogenetic distance because the proposed combination. Applicants’ argument is not persuasive because they are all fish.
Applicants cite Li (2019) and argue the gene that controls red coloration in tilapia maps to slc45a1. Applicants’ argument is not persuasive because Edvardson used salmon which are red and made them albino.
Applicants argue Edvardson taught away from homozygosity because of long breeding cycles in salmon. Applicants’ argument is not persuasive. Just because it might take time is inadequate to prevent those of skill from pursuing it.
Conclusion
No claim is 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.
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Michael C. Wilson
/MICHAEL C WILSON/
Primary Examiner, Art Unit 1638