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 .
This Office Action is in reply to Applicants’ correspondence of 10/22/2025.
Applicants’ remarks and amendments have been fully and carefully considered but are not found to be sufficient to put the application in condition for allowance. Any new grounds of rejection presented in this Office Action are necessitated by Applicants’ amendments. Any rejections or objections not reiterated herein have been withdrawn in light of the amendments to the claims or as discussed in this Office Action.
This Action is made FINAL.
Please Note: The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Election/Restrictions
In the reply filed on 05/02/2025 Applicants elected, without traverse, the invention of Group I (claims 1-16 drawn to methods), and the particular combination of probes that is SEQ ID NOs: 3, 4, 5, 6 and 7 (relevant to claim 8). The species election requirement was withdrawn (as set forth on page 2 of the Office Action of 05/22/2025) and claim 9 is examined as part of the elected invention.
Withdrawn Claim Rejections - 35 USC § 112 - Indefiniteness
Rejections of claims made under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as set forth on pages 2-4 of the Office Action of 05/22/2025, which are not reiterated below, as withdrawn in light of the amendments to the claims.
New Claim Rejections - 35 USC § 112 – Indefiniteness
Necessitated by Claim Amendments
Claims 1-14, 16, and 22-24 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.
Claims 1-14, 16, and 22-24 are unclear over the probes intended to be required and encompassed by the claimed methods. Claim 1 recites “a plurality of fluorescently-labeled oligonucleotide probes … wherein at least two of the probes each target a distinct genetic variation if present within the target nucleic acid sequence, and wherein at least one of the probes is a control probe targeting a control region present within the target nucleic acid sequence”. In this recitation is unclear if the “control probe targeting a control region” is intended to be one of the “at least two of the probes each target a distinct genetic variation if present within the target nucleic acid sequence”. Where claim 1 further recites “detecting a fluorescent signal from the control probe and a fluorescent signal from one or more of the at least two probes” it would appear that the total probes of the methods require at least three probes (i.e.: two probes for two different loci that may each have some particular content, and one probe that is used as a control). But this aspect of the claims is unclear where claim 2 depends form claim 1 and recites “wherein the plurality of fluorescently-labeled oligonucleotide probes is between about 2 ….” because if the total probes of the methods require at least three probes, then it would seem to be impossible to have any less than 3 fluorescently-labeled oligonucleotide probes.
Claims 5 and 22 are unclear over the limitations “the control region is a conserved region within the target nucleic acid sequence” (as recited in claim 5) “the control region is conserved and does not comprise a genetic variation” (as recited in claim 22). It is unclear if the limitation requires only that the particular probe in fact detects its complementary sequence in the sample of the methods, or if the limitation is intended to require that in some population of possible nucleic acid samples the target sequence is present without any alterations. This aspect of the claims is unclear where, for example, the specification (p.3) sets forth that:
SEQ ID NO: 4: fluorescent nucleotide probe with the HEX fluorophore at the 5' end and the IowaBlack FQ quencher at the 3' end that detects a constant region of the SARS-CoV- 2 spike protein.
But the sequence of the probe hybridizes to the nucleic acid sequence that encodes the amino acid at positions 489-495 of the SARS-CoV-2 spike protein (i.e.: YFPLQSY), where this region is not universally conserved among SARS-CoV-2 genomes. For example Najar et al (2023) provides a review of mutations in SARSCoV-2 and includes variants in the spike protein at amino acid positions 490 and 493.
Withdrawn Claim Rejections - 35 USC § 102 / 103
The rejections of claims made under 35 USC 102 and 103, as set forth on pages 4-9 of the Office Action of 05/22/2025, are withdrawn in light of the amendments to the claims.
New Claim Rejections - 35 USC § 102
Necessitated by Claim Amendments
Claim(s) 1-3, 5, 11-14, 22, 23 is/are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Huang et al (2011) (cited on the 892 of 05/22/2025).
Relevant to the rejected claims, Huang et al teaches fluorescence melting curve analysis for the identification of multiple mutations in an amplicon (e.g.: p.2 - Multicolor FMCA for Mutation Identification; Figure 4), and teaches a plurality of fluorescently labeled probes. Huang et al teaches contacting a nucleic acid with a pair of primers that amplify a portion of a nucleic acid that is about 60 bp to about 5000bp (e.g.: F1 and R1 in Fig 4A, where the entire Pol/RT fragment encodes 344 amino acids and thus the entire coding fragment is 1032 bases, and the primers amplify a portion of that coding fragment, relevant to claim 3) and three fluorescently-labeled (relevant to claims 2, 12 and 13) probes that each target an HBV locus (relevant to claim 11) and have distinguishable labels (relevant to claim 14) of non-variant and variant detection in particular samples (relevant to claim 22). Huang et al teaches detecting the fluorescence of probes annealed to targets nucleic acid sequences ((e.g.: Figure 4B), and provides the results from the analysis of multiple samples (relevant to claim 10), and teaches the detection of sequences that were generated separately in plasmid by PCR-mediated in vitro mutagenesis prior to analysis (p.7 - Oligonucleotides, Clinical Samples and Plasmids) thus providing for isolation of nucleic acids prior to analysis (relevant to claim 23).
With regard to the limitation of claim 1 which recites “detecting a fluorescent signal from the control probe … thereby detecting the target nucleic acid sequence”, the presence of any known content in the sample nucleic acid satisfies this limitation. For example, where the prior art provides a target with N at 236 (i.e.: wild type at 236) and M at 180 and V at 204 (i.e.: variants at 180 and 204) (see for example bottom left window of panel B in Fig 4, the reference is teaching the detection of a control probe (i.e. the FAM labeled probe) and the detection of the presence of two variants (i.e.: the ROX and TET labeled probes).
Response to Remarks
Applicants have traversed the rejections of claims as set forth in the Office Action of 05/22/2025. In so far as the new rejection set forth above in the instant Office Action applies the same reference (i.e.: Huang et al (2011)) to the amended claims, it is relevant to address Applicants traversal of the previous rejection as provided on pages 7-8 of the Remarks of 10/22/2025. Applicants have argued
Huang fails to teach all elements of claim 1 arranged as in the claims because Huang does not teach "wherein at least one of the probes is a control probe targeting a control region within the target nucleic acid sequence." Instead, Huang teaches: "The wild-type samples were always used as positive controls and water was used as no-template control."
This argument is not found to be persuasive to withdraw the rejection as set forth in the instant Office Action. The argument is not persuasive because in the context of the claims the functional requirement of the control probe is that it is “targeting a control region present within the target nucleic acid sequence” and the practical step of the claims related to the control probe is “detecting a fluorescent signal from the control probe … thereby detecting the target nucleic acid sequence”. While it is noted that limitations related to the control region are addressed in this Office Action under 35 USC 112(b) as indefinite, when considering the prior art, the only concern is whether or not the prior art includes a probe that provides a detectable fluorescence indicative of the presence of the target nucleic acid (i.e.: a control probe), as well as additional probes with detectable fluorescence indicative of the presence of some other particular genetic content in the target nucleic acid (i.e.: the probes of the claims that “each target a distinct genetic variation if present within the target nucleic acid sequence”). In the instant case the, where the rejected claims do not recite or require any particular structures (i.e.: defined polynucleotide sequences) of the probes or primers of the claims, the methods of the cited prior art meet the limitations of the claims.
New Claim Rejections - 35 USC § 103
Necessitated by Claim Amendments
Claim(s) 1-3, 4, 5, 6, 11-14, 22, 23 and 24 is/are rejected under 35 U.S.C. 103 as being unpatentable over by Huang et al (2011) (cited on the 892 of 05/22/2025) in view of Rowan et al (available online 10 May 2021).
The teachings of Huang et al are applied to the instantly rejected claims as they were previously applied in the rejection of claims 1-3, 5, 11-14, 22, 23 under 35 USC 102 above. Huang et al teaches methods of detecting particular nucleic acids in a sample using a plurality of fluorescently labeled probes. Further relevant to the instant rejection, Huang et al teaches amplification reactions that include dNTPs (e.g.: p.8 - Optimization of PCR Conditions for Probe-based FMCA).
Huang et al does not exemplify method that includer polymerase with 5’-3’ exonuclease activity, a buffer with a pH adapted for the polymerase activity of the polymerase, or reverse transcription with a reverse transcriptase (relevant to claim 4, 6 and 24). However, such elements in the context of nucleic acid detection were known in the art and are taught by Rowan et al.
Relevant to the rejected claims, Rowan teaches the detection of target nucleic acids in a sample using RT-qPCR that includes reverse transcribing RNA (i.e.: contacting sample RNA with a reverse transcriptase), and detection of a FAM-labelled hydrolysis probe (relevant to the claims as they require a polymerase with 5’-3’ exonuclease activity) via PCR using TaqMan Fast virus one step Master Mix (relevant to the claims as they require a buffer adapted to a polymerase).
It would have been prima facie obvious to someone with ordinary skill in the relevant art before the effective filing date of the rejected claims to have used the RT-qPCR methods of Rowan et al, reverse transcription and detection using hydrolysis probes via a polymerase with 5’-3’ exonuclease activity, to detect the genetic content in the HBV POL/RT region as taught by Huang et al. The skilled artisan would have been motivated to use the method of Rowan et al based on the expressed teachings of Rowan et al that such methods achieve sensitive detection of target nucleic acids. The skilled artisan would have had a reasonable expectation of success where Huang et al teaches probe sequences that detect target nucleic acid sequences and include fluorophores and quenchers (e.g.: Fig 1), and Rowan et al teaches the detection of target nucleic acid sequences using hydrolysis probes that include fluorophores and quenchers (e.g.: Table 1). The skilled artisan would recognize that where the target sequence of Huang et al is in a protein coding sequence, the detection of the content in an RNA transcript using reverse transcription as taught by Rowan et al would be an alternative target for identification of expressed variant content.
Claim(s) 7 and 16 is/are rejected under 35 U.S.C. 103 as being unpatentable over Huang et al (2011) (cited on the 892 of 05/22/2025) in view of Rowan et al (available online 10 May 2021), as applied to claims 1-3, 4, 5, 6, 11-14, 22, 23 and 24 above, further in view of Jakhmola et al (Published online March 20, 2021).
The teachings of Huang et al in view of Rowan et al are applied to the instantly rejected claims as they were previously applied in the rejection of claims 1-3, 4, 5, 6, 11-14, 22, 23 and 24 above. Huang et al in view of Rowan et al renders obvious the detection of genetic content in a target nucleic acid using a plurality of fluorescently labeled probes (as taught by Huang et al) via RT-qPCR that includes reverse transcribing RNA, and detection of a labeled hydrolysis probe via PCR with polymerase with 5’-3’ exonuclease activity (as taught by Rowan et al). Further relevant to the instantly claimed methods, Rowan et al exemplifies the detection of a target nucleic acid that is the envelope gene for Sars-CoV-2 using primers and a probe directed to a conserved portion of the E gene (e.g.: p.6, left col: the Charite E primers and probe are extremely sensitive and can detect >99.9 % of variants deposited in GISAID at the time of writing).
Huang et al in view of Rowan et al does not provide for the detection of content at a plurality of regions in the Sars-CoV-2 E gene. However, the existence of multiple variants of the E gene in the Sars-CoV-2 family of viruses was known in the prior art and is taught by Jakhmola et al.
Jakhmola et al teaches that analysis of Sars-CoV-2 genome sequences from publicly available databases (e.g.: p.2 - Sequence retrieval and multiple sequence alignment (MSA)) and the identification of mutations in the E gene in the genomes (e.g.: p.3 - Multiple sequence alignment of SARS-CoV-2 E, M and S proteins revealed unique mutations). The reference disclose specific mutations in the E gene associated with SARS-CoV-2 isolated at specific geographic locations at particular times (e.g.: Figure 2).
It would have been prima facie obvious to someone with ordinary skill in the relevant art before the effective filing date of the rejected claims to have applied the genetic variant detection rendered obvious by Huang et al in view of Rowan et al to the detection of SARS-CoV-2 E gene variants as taught by Jakhmola et al. The skilled artisan would have been motivated to detect the E gene variants taught by Jakhmola et al based on the expressed teachings of Jakhmola et al that variants in the E gene may have functional consequences with regard to protein function. The skilled artisan would have a reasonable expectation of success where Jakhmola et al sequences relevant to SARS-CoV-2 can be accessed from databases.
Conclusion
No claim is allowed.
Claims 8 and 9 are free of the prior art as set forth on page 10 of the Office Action of 05/22/2025.
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 STEPHEN THOMAS KAPUSHOC whose telephone number is (571)272-3312. The examiner can normally be reached M-F, 8am-5pm.
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Stephen Kapushoc
Primary Examiner
Art Unit 1683
/STEPHEN T KAPUSHOC/Primary Examiner, Art Unit 1683