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 .
Status of Claims
Applicant's amendment filed 02/11/2026 is acknowledged. Claims 1-3, 6-10, and 12-14 have been amended. Claims 16-21 have been added. Claim 15 has been cancelled. Claims 1-14 and 16-21 are pending in the instant application and the subject of this final office action.
All of the amendments and arguments have been reviewed and considered. Any rejections or objections not reiterated herein have been withdrawn in light of amendments to the claims or as discussed in this office action.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
New Ground(s) of Rejections/Status of Previous Rejections
Status of Prior Rejections/Objections:
The objections to the specification and claim 9 are withdrawn in view of the respective amendments.
The 112(b) rejections of claim(s) 12-13 and 15 are withdrawn in view of the amendments to or cancellation of the claims.
The prior art rejection(s) under 35 USC 103 directed to claim(s) 1-14 as being unpatentable over Georgiou in view of Uphoff, as evidenced by Tanner and Hopert are maintained. See new 103 rejections of new claims 16-21 over the same.
The 101 rejection of claims 1-13 and 15 is withdrawn in view of Applicants’ amendments and arguments.
The double patenting rejections over copending applications 18009498 and 18009142 are maintained.
The new ground(s) of rejections were necessitated by applicant’s amendment of the claims.
Information Disclosure Statement
The Information Disclosure Statements filed February 11, 2026 and March 6, 2026 have been considered.
Claim Objections
Claims 19 and 20 are objected to because of the following informalities:
Claims 19 and 20 recite, in line 1 of each claim, “derived from”; this should be changed to “obtained from”.
Appropriate correction is required.
Claim Interpretation
In evaluating the patentability of the claims presented in this application, claim terms have been given their broadest reasonable interpretation (BRI) consistent with the specification, as understood by one of ordinary skill in the art, as outlined in MPEP 2111.
Regarding claim 1, the claim recites “no outer forward primer (F3)”. The specification recites that “The term ‘F3’, as used herein, refers to the outer forward primer of a primer set” (pg. 7, para 1) and that “The primer set according to the present invention is used for amplification of nucleic acids, that is for a LAMP analysis or a RT-LAMP analysis” (pg. 5, para 2). Therefore, the “outer forward primer (F3)” has been interpreted to be an outer forward primer/F3 primer useful in a LAMP or RT-LAMP amplification, consistent with the art accepted definition of an “F3 primer”.
Regarding claim 19, the claim recites “the … sequence is derived from … a cytoplasm … of the bacterium of the Mollicutes class”. Prokaryotic Mollicutes lack membrane-bound organelles. Therefore, the cytoplasm is interpreted broadly to encompass all portions of the bacteria enclosed by the cytoplasmic membrane, i.e., inner structures excluding cell/cytoplasmic/plasma membrane(s) and including, for example, the nucleoid.
Claim Rejections - 35 USC § 112(b)
Claims 12 and 17-19 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.
Regarding claim 12, the claim recites the limitation "the elongated strand" in line 3. There is insufficient antecedent basis for this limitation in the claim.
Regarding claim 17, the claim recites “wherein five DNA primers are added to the sample as follows: … FIP…BIP…LPF…LPB…and…B3”. It is unclear whether this limitation requires that exactly and only these five DNA primers be added to the sample in step (b) such that the addition of any further primers to the sample (in step (b) or other unclaimed steps) would be outside of the scope of this claim or if the limitation merely specifies the identity of five particular DNA primers of any of at least five DNA primers that may be added to the sample.
Regarding claim 18, the claim recites “the culture” in steps (e) and (f). There is insufficient antecedent basis for this limitation in the claim.
Regarding claim 19, the claim recites “the … nucleic acid sequence is derived from a nucleus … a mitochondria … of a bacterium of the Mollicutes class”. It is unclear how to derive sequences from nuclei or mitochondria of bacteria that are prokaryotes and lack these organelles.
Claim Rejections - 35 USC § 112(d)
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.
Claim 19 is 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. 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.
Regarding claim 19, the claim recites “the … sequence is derived from … a ribosome… a cytoplasm, or a plasmid of the bacterium of the Mollicutes class”. As discussed in part in the interpretation, prokaryotic Mollicutes cells consist of a plasma membrane and a cytoplasm comprising genomic DNA, ribosomes, etc. As the sources of nucleic acids of prokaryotes are within the cytoplasm and even extracellular vesicles would be interpreted broadly to be derived from “a cytoplasm”, the limitation that “the pre-determined nucleic acid sequence be derived from … a cytoplasm … of a bacterium of the Mollicutes class” fails to limit the claim beyond “a pre-determined nucleic acid sequence of a bacterium of the Mollicutes class”.
For this reason, claim 19 fails to further limit claim 4 from which it depends and, therefore, does not comply with the requirements of 112(d).
Claim Rejections - 35 USC § 103
Claim(s) 1-14 and 16-21 is/are rejected under 35 U.S.C. 103 as being unpatentable over Georgiou (WO 2019/234251 A1; published 12/12/2019) in view of Uphoff (Uphoff CC, et al. Treatment of mycoplasma contamination in cell cultures with Plasmocin. J Biomed Biotechnol. 2012;2012:267678. Epub 2012 Oct 2.) as evidenced by Tanner (New England Biolabs, Inc. Isothermal Amplification [Internet]. Ipswich (MA): New England Biolabs, Inc.; 2015 Apr [cited 2025 Aug 26]. Available from: https://www.neb-online.de/wp-content/uploads/2015/04/NEB_isothermal_amp.pdf) and Hopert (Hopert A, et al. Specificity and sensitivity of polymerase chain reaction (PCR) in comparison with other methods for the detection of mycoplasma contamination in cell lines. J Immunol Methods. 1993 Aug 26;164(1):91-100).
Regarding claims 1, 4-8, 17, 19, and 21, Georgiou teaches a method for determining presence of a pre-determined nucleic acid sequence in a sample (pg. 47), the method comprising the steps of:
(a) adding an enzyme to the sample to be analysed for the presence of the pre-determined nucleic acid sequence (pg. 47, lines 5-8), wherein the enzymes provide activities of RNA- and/or DNA-dependent DNA polymerase activity and strand-displacement activity (pg. 9, para 5-6; pg. 2, para 9);
(b) adding at least five DNA primers to the sample to be analysed for the presence of the pre-determined nucleic acid sequence, wherein at least one DNA primer comprises a sequence hybridisable to the nucleic acid sequence and at least one DNA primer comprises a sequence hybridisable to the DNA sequence reverse-complementary to the nucleic acid sequence (pg. 47: (iv): FIP, (v): BIP, (vi): forward blocking, (vii): backward blocking; pg. 23, lines 10-11: forward loop primer and backward loop primer; pg. 22, lines 22-23; pg. 22, lines 36-37)
(c) incubating the sample resulting from steps (a) and (b) at a fixed temperature (pg. 47, (a): “amplifying under isothermal conditions” [i.e., a fixed temperature]);
(d) determining whether an elongated DNA sequence is present in the sample, wherein presence of the elongated DNA sequence in the sample is indicative of the presence of the pre-determined nucleic acid sequence in the sample (pg. 47, (b))
Georgiou teaches a polymerase with strand-displacement activity at least by teaching that the DNA polymerase when the nucleic acid sequence is DNA (instant claim 4 and 8) may be Bst 2.0 DNA polymerase (pg. 9, para 5), evidenced by Tanner. While Georgiou does not explicitly teach that the polymerase itself has strand-displacement activity, it is inherently taught because Bst 2.0 has strand-displacement activity, evidenced by Tanner (pg. 4, para 1: “Bst 2.0 DNA polymerase possesses … strong strand-displacement activity”).
Georgiou teaches that if the nucleic acid sequence is RNA, the method may also comprise reverse transcription of the RNA to produce cDNA (pg. 9, lines 36-40; instant claims 4 and 7).
Georgiou teaches an embodiment without F3 (pg. 22, lines 22-23: “The reaction mixture may further comprise … F3 … and/or … B3 …” (emphasis added); pg. 22, lines 36-37: “The present inventors have determined that detectable amplification of the nucleic acid can occur in the absence of F3 and/or B3”). Georgiou teaches [traditional] LAMP uses four to six primers in the reaction (pg. 2, line 9), and that the efficiency of the reaction is enhanced by the addition of B3 (pg. 23, lines 6-7) and LF and LB (pg. 23, line 22).
While Georgiou does teach that the species the method is intended to target include bacteria (pg. 48, para 2), Georgiou does not teach:
that the pre-determined nucleic acid sequence is from a bacterium of the Mollicutes
class (claim 1);
particular genera or species (claims 5-6);
that the target is a 16S or 23S rRNA sequence (claims 7-8);
the samples of claim 9; and
decontaminating using an antibiotic (claims 13-14 and 18).
Uphoff teaches that cell culture has become an important research tool, wherein one of the major problems of the cell culture field is infection with mycoplasma, which can affect every cellular parameter, often leading to experimental artefacts and spurious results (pg. 1, Introduction, para 1).
Uphoff teaches verification of mycoplasma contamination by targeting the 16S rRNA gene of bacteria including M. arginini [i.e., Mycoplasma arginini], M. fermentas, A. laidlawii [i.e., Acholeplasma laidlawii], M. pneumoniae, and M. orale (instant claims 5-6), evidenced by Hopert. While Uphoff does not explicitly teach targeting the 16S rRNA of those bacteria species, it is inherently taught because Uphoff utilizes the methods of Hopert (Uphoff: pg. 2, 2.2. Mycoplasma Detection and Species Identification by PCR, para 1; Uphoff: Table 1), as evidenced by Hopert (Hopert: Abstract; Table 1; instant claim 7-8).
Uphoff teaches that the advent of specific antimycoplasma antibiotics together with the availability of the most sensitive and robust detection method represent efficient methods to achieve mycoplasma-free cultures (pg. 6, Discussion, para 1), wherein it is mandatory to document the [treatment] outcome with sensitive assays in order to verify complete and permanent freedom from mycoplasma (pg. 7, col 2, para 2).
Therefore, it would have been obvious to one of ordinary skill in the art before the effective of filing date of the claimed invention to have utilized the 16S rRNA gene of the above Mollicutes species as a target, as taught by Uphoff as evidenced by Hopert, in the method of Georgiou as evidenced by Tanner utilizing either RNA or DNA as a target nucleic acid, motivated by the desire enable verification of both the infection of cell cultures and the successful treatment to address the problematic infection of research cell cultures, as taught by Uphoff as evidenced by Hopert. There would have been a strong expectation for success as both are directed to identifying and treating infectious bacteria using amplification means, wherein one represents the application of a known technique to a known method.
It further would have been oblivious to one of ordinary skill in the art before the effective filing date of the claimed invention to add LF [instant LPF] and LB [instant LPB] (instant claim 2) and (instant claim 17)/or B3 (instant claim 3) in the method of determining the presence of a sequence in a sample Georgiou evidenced by Tanner, motivated by the desire to increase the efficiency the reaction, as taught by Georgiou.
There would be a strong expectation for success as the methods fall within the scope of Georgiou and are a combination of prior are elements according to known methods to yield predictable results.
Further, while Georgiou teaches embodiments with FIP, BIP, FB, BB, LF, LB, and B3 (i.e., at least 5 primers; instant claim 17), it is further noted that the total number of primers utilized represents a matter for efficiency/optimization, as taught by Georgiou (e.g., typical number of LAMP primers, addition of LB and LF and/or B3).
It is noted that the courts have found that “where 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). See MPEP 2144.05 II. Thus, the claimed range of the number of primers merely represents routine optimization of the values of the cited prior art.
Interpreted under the broadest reasonable interpretation, claim 17 requires the five DNA primers FIP, BIP, LPF, LPB, and B3, which are taught in an embodiment of Georgiou. The
As noted above regarding the interpretation of claim 19, in teaching the combination of steps directed to a pre-determined sequence of a bacterium of the Mollicutes class of claim 1, the limitation of claim 19 would be met.
Regarding claim 2, in the method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert, as cited in the rejection of claim 1, Georgiou teaches primers including FIP and BIP in the method and an embodiment in which LF [instant LPF] and LB [instant LPB] are added to improve efficiency. The same motivations and expectations for success as above apply.
Regarding claim 3, in the method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert, as cited in the rejection of claim 1, Georgiou teaches B3 is added to improve efficiency. The same motivations and expectations for success as above apply.
Regarding claims 9 and 20, in the method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert, Uphoff teaches sampling cell cultures (pg. 2, 2.2. Mycoplasma Detection and Species Identification by PCR, para 1).
Georgiou also teaches samples including plant matter, animal matter, soil, a clinical and sample from a patient [i.e., a living body] (pg. 24, para 1-2).
Regarding claim 10, in the method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert, Georgiou teaches that the temperature for the isothermal conditions may be around 50 C to 70 C (pg. 6, line 8).
Regarding claim 11, in the method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert, Georgiou teaches the amplifying duration may be around 1 to 150 minutes and ranges in between (pg. 6, para 2).
Regarding claims 12 and 16, in the method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert, Georgiou teaches detection of amplification products [i.e., double-stranded elongated DNA sequences/molecules] by turbidity and nucleic acid intercalating agents (pg. 7, lines 7-8).
Regarding claims 13, 14, and 18, in the method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert, Uphoff further teaches decontaminating a cell culture infected by a bacterium of the Mollicutes class by administering to the culture antimycoplasma antibiotic treatment protocols including BM-Cyclin (pg. 3, 2.3. Antimycoplasma Treatment and 2.4. Antimycoplasma Antibiotics).
Uphoff teaches that BM-Cyclin had the lowest resistance rate and highest cure rate of the three primary classes tested (Fig. 2).
Therefore, it further would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have treated the cultures that have been identified as infected using the combined method of Georgiou in view of Uphoff as evidenced by Tanner and Hopert with an antimyoplasma antibiotic, motivated by the desire to address the problematic infection of research cell cultures, as taught by Uphoff. It further would have been obvious to the POSITA before the EFD to have chosen BM-Cyclin, motivated by the low resistance rate and high cure rates identified. There would have been a strong expectation for success as the methods are directed to identifying and treating infectious bacteria using amplification means wherein such an improvement represents the application of a known technique to a known method.
Double Patenting
Claims 1-14 and 16-21 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-6 and 8-10 and 16 of copending Application No. 18/009,498 in view of Uphoff (Uphoff CC, et al. Treatment of mycoplasma contamination in cell cultures with Plasmocin. J Biomed Biotechnol. 2012;2012:267678. Epub 2012 Oct 2.) as evidenced by Hopert (Hopert A, et al. Specificity and sensitivity of polymerase chain reaction (PCR) in comparison with other methods for the detection of mycoplasma contamination in cell lines. J Immunol Methods. 1993 Aug 26;164(1):91-100). This is a provisional nonstatutory double patenting rejection.
Both sets of claims are claims are directed to a method of determining a presence of a pre-determined nucleic acid sequence in a sample comprising the steps of adding an RNA/DNA-dependent DNA polymerase with strand-displacement activity and at least five primers, incubating at a fixed sample, and determining whether an elongated DNA sequence is present in the sample, wherein the elongated DNA sequence is indicative of the pre-determined sequence in the sample and wherein no F3 primer is used, wherein the sample may be obtained from an animal from an animal.
‘498 claim 6 recites that the pathogen may be a bacterium.
‘498 claim 16 recites that the determining may use a turbidity measurement.
Any additional limitations of the ‘498 claims are encompassed by the open claim language “comprising” found in the instant claims.
‘498 claim 3 teaches the set of five DNA primers claimed in instant claim 17.
‘498 fails to teach specific species/genera of the Mollicutes class, a 16S or 23S gene target, a sample from cell culture or similar, or decontaminating an infected cell/tissue culture with an antibiotic including BM Cyclin.
Uphoff, as evidenced by Hopert, rectifies this. The teachings of Uphoff, as evidenced by Hopert, from the rejections of instant claim 1, 6-9, and 13-14, 18, 20, 21 above are incorporated from above.
Uphoff further teaches that the cell lines of the method encompass animal cell lines from various tissue/cell types [i.e., were originally obtained from animal subjects] (Table 2). It is noted that the method of ‘498 requires that the sample be obtained from the animal subject and that the method states that is “comprising” the steps listed. Therefore, it has been interpreted to encompass the establishment of a cell culture from the animal sample.
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine ‘498 with the 16S method of identifying the Mollicutes species and treating with BM Cyclin of Uphoff as evidenced by Hopert, motivated by the desire enable verification of both the infection of cell cultures and the successful treatment to address the problematic infection of research cell cultures, especially with the low resistance rate/high cure rate of BM Cyclin, as taught by Uphoff . There would have been a strong expectation for success as the methods are directed to identifying and treating infectious bacteria using amplification means wherein such an improvement represents the application of a known technique to a known method.
Claims 1-14 and 16-21 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claim 1-10 and 15-17 of copending Application No. 18/009,142 in view of Uphoff (Uphoff CC, et al. Treatment of mycoplasma contamination in cell cultures with Plasmocin. J Biomed Biotechnol. 2012;2012:267678. Epub 2012 Oct 2.) as evidenced by Hopert (Hopert A, et al. Specificity and sensitivity of polymerase chain reaction (PCR) in comparison with other methods for the detection of mycoplasma contamination in cell lines. J Immunol Methods. 1993 Aug 26;164(1):91-100). This is a provisional nonstatutory double patenting rejection.
Both sets of claims are claims are directed to a method of determining a presence of a pre-determined nucleic acid sequence in a sample comprising the steps of adding an RNA/DNA-dependent DNA polymerase with strand-displacement activity and at least five primers, incubating at a fixed sample, and determining whether an elongated DNA sequence is present in the sample, wherein the elongated DNA sequence is indicative of the pre-determined sequence in the sample and wherein no F3 primer is used.
‘142 claim 7 recites that the pathogen may be from the genus Mycoplasma, which is part of the Mollicutes class, as evidenced by instant claim 5.
‘142 claim 15 recites the detection using an intercalating molecule or turbidity measurement.
‘142 claims 16-17 recites the set of five LAMP primers of instant claim 17.
Any additional limitations of the ‘142 claims are encompassed by the open claim language “comprising” found in the instant claims.
‘142 fails to teach specific species of the Mollicutes class, a 16S or 23S gene target, a sample from cell culture or similar, or decontaminating an infected cell/tissue culture with an antibiotic including BM Cyclin.
Uphoff, as evidenced by Hopert, rectifies this. The teachings of Uphoff, as evidenced by Hopert, from the rejections of instant claim 1, 6-9, and 13-14 are incorporated from the rejections above.
Uphoff further teaches that the cell lines of the method encompass human cell lines from various tissue/cell types [i.e., were originally obtained from human subjects] (Table 2). It is noted that the method of ‘142 requires that the sample be obtained from the human subject and that the method states that is “comprising” the steps listed. Therefore, it has been interpreted to encompass the establishment of a cell culture from the human sample.
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine ‘142 with the 16S method of identifying the Mollicutes species and treating with BM Cyclin of Uphoff as evidenced by Hopert, motivated by the desire enable verification of both the infection of cell cultures and the successful treatment to address the problematic infection of research cell cultures, especially with the low resistance rate/high cure rate of BM Cyclin, as taught by Uphoff . There would have been a strong expectation for success as the methods are directed to identifying and treating infectious bacteria using amplification means wherein such an improvement represents the application of a known technique to a known method.
Response to Arguments
Applicant's arguments filed 02/11/2026 have been fully considered but they are not persuasive.
Regarding the 103 rejections, on pgs. 9-11, Applicant argues that Georgiou teaches a method based on six sbLAMP primers and two blocking primers and that, unlike Georgiou, the instant method is not directed to detection of specific SNPs. Applicant argues that the Office has not provided a motivation for why the artisan would have removed the blocking primers of Georgiou or an adequate evidentiary basis for the finding that includes a rational connection with the evidence presented as to why the artisan would modify Georgiou in view of Uphoff, Hopert, and Tanner to remove the blocking primers to arrive at the instant claims. Applicant argues that the removal of the blocking primers would lead to inoperability of Georgiou’s method. Applicant further argues that Georgiou fails to provide experimental evidence showing that omission of F3 is possible for a five primer system or that such a five-primer system without F3 would allow efficient amplification. Applicant also argues that Georgiou does not teach the predetermined nucleic acid sequence is from the bacterium of Mollicutes class and that each Hopert, Uphoff, and Tanner fail to teach the full method.
Applicant additionally argues that the inventors have surprisingly found that a reaction in the presence of B3 but absence of F3, detection is faster and more sensitive and alleges that this is an unexpected result.
In response to the arguments about the teachings of Georgiou regarding six SNP-based primers and two blocking primers, Georgiou teaches multiple embodiments. As cited, Georgiou teaches embodiments that exclude F3 (see pg. 22, lines 22-23; pg. 22, lines 36-37 and pg. 22, lines 22-23). Further, the claims as written recite “adding at least five DNA primers” and “comprising the steps of”. Accordingly, the claims encompass adding the blocking primers of Georgiou and there is no further need to provide a motivation or factual finding for their removal or that of F3. It is noted that a motivation has been provided to add LF [instant LPF] and LB [instant LPB] and/or B3 to increase efficiency of the reaction, as taught by Georgiou and that the total number of primers is further subject to routine optimization, as taught by Georgiou. The clerical error referring to “B3” as “L3” in the paragraph discussing routine optimization has been corrected. As there is no need for a motivation for the removal of the blocking primers in the combined method, the arguments directed to the inoperability are moot.
It is noted that the features upon which applicant relies (i.e., no blocking primers) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
Likewise, regarding the arguments directed at the inoperability of Georgiou in embodiments absent F3 and the efficiency of such a method, MPEP 2121(I) recites: “When the reference relied on expressly anticipates or makes obvious all of the elements of the claimed invention, the reference is presumed to be operable. Once such a reference is found, the burden is on applicant to rebut the presumption of operability”. Georgiou expressly discusses that detectable amplification of the nucleic acid can occur in the absence of F3 and/or B3 and discusses possible mechanisms in pg. 22, para 7, spanning pg. 23. Where the applicant makes arguments directed to a level of efficiency, it is noted that the features upon which applicant relies (i.e., “efficiency relative to an amplification including F3”) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
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Regarding the allegations of unexpected results, as discussed in MPEP 716.02: the "objective evidence of nonobviousness must be commensurate in scope with the claims which the evidence is offered to support." The measured fluorescence, shown in Fig. 1, of the “5 primer” is not clearly distinguishable from the “no sample” control and the “6 primer” at 10 minutes. This raises a question as to whether genuine amplification, as opposed to background signal, has been detected at this time. Further, where clear divergence of “5 primer” and “6 primer” occur at about 15 min, the “6 primer” appears to be the more efficient with a higher relative fluoresce in a shorter amount of time, indicating that any increase in efficiency should one be shown, appears to be transient. Therefore, absent supporting data (e.g., statistical replicates, threshold values, or error bars) and given the current broad scope of the claims, the evidence of unexpected results is not persuasive.
The context of the data presented being a single replicate in a system recited to be applicable to “a low number of … sequence in a sample” (Remarks, pg. 11, para 1, citing the instant specification para [0053]) is also noted. The art teaches that LAMP has increased variation at low numbers of template molecules. Namely, Hardinge (Hardinge P, Murray JAH. Full Dynamic Range Quantification using Loop-mediated Amplification (LAMP) by Combining Analysis of Amplification Timing and Variance between Replicates at Low Copy Number. Sci Rep. 2020 Jan 22;10(1):916) teaches that real-time monitoring of LAMP reactions by at least fluorescence dye binding shows increasing variation between replicates at low copy number due to the LAMP amplification mechanism, and that this effect increases with decreasing copy number and is associated with the amplification frequency (Abstract). Hardinge teaches that the increasing variation is not linked to the use of displacement primers (Abstract).
Regarding the arguments about the intended use of the SNP detection of Georgiou, it is noted that the features upon which applicant relies (i.e., determining the presence of a Mollicutes class bacterium) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993). The claims require determining whether a double-stranded elongated DNA sequence indicative of the presence of a pre-determined sequence of a bacterium is present. Only claim 4 requires the sequence to be derived from the bacteria itself.
Further, as cited, Georgiou teaches that method for pathogen microorganisms including for bacteria (see pg. 48, para 2-3 and pg. 1, para 2). The claims, as drafted, do not require any particular target pre-determined nucleic acid sequence target that would exclude the targeting of a SNP. Indeed, single nucleotides polymorphisms within, for example, the 16S gene are widely used genetic markers, especially for species and subspecies identification, including for Mycobacterium. See, for example, Chakravorty (Chakravorty S, et al. A detailed analysis of 16S ribosomal RNA gene segments for the diagnosis of pathogenic bacteria. J Microbiol Methods. 2007 May;69(2):330-9. Epub 2007 Feb 22): “A separate analysis of three … regions … within V2 … contained the maximum SNP variations between Mycobacterium and Haemophilus species and the region spanning 205–242 contained most of the SNP variation between the Staphylococcal, Streptococcal, Haemophilus, Fusobacterium, Clostridium and Neisseria species as well as other bacterial genus analyzed” (3.3. Region V2 (nucleotides 137–242)). Indeed, the primers of Uphoff demonstrate examples of what the artisan would recognize to be likely polymorphisms within homologous sequence (Table 1). And, as discussed in the 103 rejections, the combination renders the detection using the combined method, motivated by, at least, the desire to enable verification of both the infection of cell cultures and the successful treatment to address the problematic infection of research cell cultures.
In response to applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986).
Regarding the double patenting rejections, Applicant requests that the provisional rejections be held in abeyance.
As recited in MPEP 804(I)(B)(1): “As filing a terminal disclaimer, or filing a showing that the claims subject to the rejection are patentably distinct from the reference application’s claims, is necessary for further consideration of the rejection of the claims, such a filing should not be held in abeyance”. Accordingly, the rejections are maintained and modified in view of amendments.
Conclusion
No claims are allowed.
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure.
Gong (CN 103898222 B; published 09/09/2015) teaches a detection method of a bacteria (entire document, e.g., para [0002] and claims 2-3) comprising adding Bst polymerase (e.g., claims 1-3; para [0082]); adding the DNA primers four to six of the primers F3, B3, FIP, BIP, LF, and LB (e.g., claims 1-3; para [0068-0069]); incubating at a fixed temperature (claim 2, step 4; claim 2; Example 1); and determining the presence of the target bacterial sequence via a double-stranded elongated DNA sequence using SYBR green (claim 3).
Gong teaches determining the efficiency of the reaction [i.e., optimizing] by leaving out F3, B3, or both F3 and B3 (Fig. 3; para [0068]). Gong teaches that absence of an outer primer has less of an effect on the LAMP reaction [i.e., with their single set of primers tested] than an inner primer (para [0069]). Gong also teaches optimizing the concentration of the ratio of outer primers to inner primers (para [0065]).
Gong teaches the polymerase Bst (para [0082], claim 1), i.e., an RNA- and/or DNA- dependent polymerase with strand-displacement activity.
That is to say, Gong teaches performing a LAMP for the detection of a bacteria with five primers B3, FIP, BIP, LF, and LB using a strand-displacing polymerase.
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).
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/EMMA R HOPPE/Examiner, Art Unit 1683
/NANCY J LEITH/Primary Examiner, Art Unit 1636
