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
Applicant's election with traverse of Group I (claims 1-6, 9-13 and 22-23) in the reply filed on 10/29/2025 is acknowledged. The traversal is on the ground(s) that “The Legal Standard The relevant standard for applications filed in the national stage under 35 U.S.C. § 371, is the Unity of Invention requirement under PCT rule 13.1 and 13.2. See MPEP § 802. The basic principle for Unity of Invention is that an application should relate to only one invention or, if there is more than one invention, that Applicant would have a right to include in a single application only those inventions which are so linked as to form ''a single general inventive concept." A group of inventions is considered to be linked to form a single general inventive concept where there is a technical relationship among the inventions that involves at least one common or corresponding special technical feature.
When making a lack of Unity of Invention allegation, the Examiner must (1) list the different Groups of claims and (2) explain why each Group lacks unity with each other Group (i.e., why there is no single general inventive concept) specifically describing the unique special technical feature in each Group.
A Group of inventions is considered to be linked to form a single general inventive concept where there is a technical relationship among the inventions that involves at least one common or corresponding special technical feature. See MPEP 1893.03(d).
Analysis
Claim 1 as amended, and claims 2-6, 9-19, and 21-23 dependent thereof, are based at least on Applicant's development of a specific one-step RT-PCR reagent composition containing
wild-type Moloney Murine Leukemia Virus reverse transcriptase (MMLV-RT) and optionally, a Thermus aquaticus DNA polymerase (Tag Pol) that is not Platinum Tag polymerase. As specified in claim 2, the MMLV-RT and Tag Pol enzymes are present in the RT-PCR reagent composition in a ratio of 2:1 (i.e., 2 units of MMLV-RT to 1 unit of Tag Pol), and formulated in a buffer for simultaneous reverse transcription and PCR amplification (Application, pp. 57, lines 9-34; pp. 64, lines 13-26).
Applicant demonstrated that a one-step RT-PCR reagent composition containing wild-type MMLV-RT and a Tag Pol that is not Platinum Tag polymerase, present in a ratio of 2:1, allows effective one-step RT-PCR without loss of PCR activity due to reverse transcriptase inhibition (see Application, pp. 57, lines 9-34; pp. 64, lines 13-26; Figures 3C-3E). In particular, Applicant experimentally confirmed that MMLV-RT at amounts conventionally used for cDNA
synthesis (e.g., >100 ng) completely inhibits Tag polymerase activity, even at elevated Tag Pol units, and that only upon identifying and implementing the claimed 2:1 MMLV-RT:Taq Pol ratio could one-step RT-PCR be successfully performed (see Application as filed, at least page 57, lines 10-31; Figures 2F and 3C).
The wild-type MMLV-RT and Tag Pol present in a defined 2:1 ratio in the reagent composition thus constitutes at least a technical feature that links the claims of Groups I-IV. Specifically, the claims of Group I define an RT-PCR reagent composition containing wild-type MMLV-RT and a Tag polymerase that is not Platinum Tag polymerase, wherein the enzymes are present in a defined 2:1 ratio and in a buffer effective for one-step RT-PCR. The claims of Groups II-IV define methods of performing one-step RT-PCR (Group II), detecting the presence of a nucleic acid in a sample (Group Ill), and diagnosing a subject for infection with a virus (Group IV), respectively, each of which directly (i.e., Groups II and Ill) or indirectly (Group IV) requires contacting a sample with the composition of claim 1. Therefore, the specific enzyme pairing of wild-type MMLV-RT and non-Platinum Tag polymerase in the defined 2:1 ratio constitutes the shared technical feature linking the Group I claims (RT-PCR reagent compositions) and the Group II-IV claims (methods of using the same compositions to perform one-step RT-PCR, detect nucleic acids, and diagnose viral infection respectively).
Corman is directed to the detection of SARS-CoV-2 RNA using a commercially available one-step RT-PCR kit, namely the Superscript™ III One-Step RT-PCR System with Platinum Tag Polymerase (see Corman, pp. 2, col. 2, last para.). Corman relies entirely on the manufacturer-provided formulation and instructions of that commercial kit and does not disclose or suggest modifying or replacing the RT or Platinum Tag Pol. Corman provides no teaching or guidance regarding the use of wild-type MMLV-RT in combination with a Tag polymerase that is not Platinum Tag polymerase, nor does it disclose or contemplate the ratio of 2:1 required by claim 1 as amended.
Because Corman relies exclusively on a commercial Platinum Tag-based system, it does not address, let alone resolve, the inhibitory effect of wild-type MMLV-RT on Tag polymerase activity. As discussed in the Application as filed, per manufacturer instructions, the recommended amount of reverse transcriptase for cDNA synthesis using random hexamer or dT primers is approximately 200 units (1 µg), (Application, pp. 57, lines 9-34; pp. 64, lines 13-26). However, Applicant demonstrated that C-His/Strep MMLV-RT at amounts of 100 ng or more completely inhibits Tag polymerase activity, even when as much as 20 units of His-Tag Pol are present (see Application, pp. 57, lines 9-34; pp. 64, lines 13-26). Corman contains no recognition of this inhibition problem and no teaching as to how such inhibition could be overcome when using a wild-type MMLV-RT and a Tag polymerase that is not Platinum Tag Pol.
Furthermore, the present application and claim 1 as amended, are not focused on the enzymes used in the commercial Superscript™ III Platinum™ kit relied upon by Corman. Rather, claim 1 as amended expressly requires that the Tag polymerase is not Platinum Tag Polymerase (see Application as filed, pg. 63, lines 7-9), and is directed to a specific, experimentally derived combination of wild-type MMLV-RT and non-Platinum Tag polymerase ratio of 2:1, formulated in a buffer effective for one-step RT-PCR.
For at least the reasons stated above, the technical feature that links all of the claims is not found in Corman.
Rejoinder of claims 14-19 and 21, and examination of claims 1-6, 9-19, and 21-23 is respectfully requested.”
This is not found persuasive because the common technical feature does not make a contribution over the prior art in view of Corman et al. as well as the prior art of the 35 U.S.C. 103 rejections below.
The requirement is still deemed proper and is therefore made FINAL.
Claims Status
Claims 1-6, 9-19, and 21-23 are pending.
Claims 14-19 and 21 are withdrawn.
Claims 7-8 and 20 are canceled.
Claims 1-6, 9-13 and 22-23 are currently under examination.
Priority
This application claims the benefit of and priority to U.S. Provisional Application No. 63/055,250, filed July 22, 2020, and U.S. Provisional Application No. 63/214,738, filed June 24, 2021. Receipt is acknowledged of certified copies of papers required by 37 CFR 1.55. Accordingly, the priority date of claims 1-5 and 9-13, filed on March 2, 2026, is determined to be July 22, 2020. The priority date of claim 6, filed on March 2, 2026, is determined to be June 24, 2021, the filing date of the provisional application 63/214,738. The priority date of claims 22-23, filed on March 2, 2026, is determined to be January 23, 2023, the filing date of the original claims filed in the instant application.
Nucleotide and/or Amino Acid Sequence Disclosures
Specific deficiency - This application fails to comply with the requirements of 37 CFR 1.821 - 1.825. This application contains a “Sequence Listing” as a PDF file (37 CFR 1.821(c)(2)) or as physical sheets of paper (37 CFR 1.821(c)(3)). A copy of the "Sequence Listing" in computer readable form (CRF) has been submitted; however, the content of the CRF does not comply with one or more of the requirements of 37 CFR 1.822 through 1.824, as indicated in the "Error Report" that indicates the "Sequence Listing" could not be accepted. Refer to attachment or document "Computer Readable Form (CRF) for Sequence Listing – Defective" dated 04/07/2026.
Reviewer’s comment: SEQ ID NOs: 1-29 contain an insufficient response for numeric identifier <223>. The intent of numeric identifier <223> is to explain the source of the "Artificial sequence." The current reply, "Artificial sequence," does not adequately explain the source of the genetic material, and simply repeats information already contained in the <213> field. The reply in the <223> field explaining an artificial sequence must provide some information that goes beyond what is already contained in the <213> field. If the sequence is put together from several organisms, please list those organisms. If the sequence is made in the laboratory, please indicate that the sequence is synthesized.(Pg. 1, Sequence listing in computer readable format is defective (CRFD) submitted 04/07/2026)
Required response – Applicant must provide:
A replacement "Sequence Listing" part of the disclosure, as described above in item 1); together with
An amendment specifically directing its entry into the application in accordance with 37 CFR 1.825(b)(2);
A statement that the "Sequence Listing" includes no new matter as required by 37 CFR 1.825(b)(5); and
A statement that indicates support for the amendment in the application, as filed, as required by 37 CFR 1.825(b)(4).
If the replacement "Sequence Listing" part of the disclosure is submitted according to item 1) a) or b) above, Applicant must also provide:
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3), and 1.125 inserting the required incorporation-by-reference paragraph, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter and
An amendment to the specification to remove the “Sequence Listing previously submitted as a PDF file (37 CFR 1.821(c)(2)) or as physical sheets of paper (37 CFR 1.821(c)(3))
If the replacement "Sequence Listing" part of the disclosure is submitted according to item 1) c) or d) above, Applicant must also provide:
A CRF in accordance with 1.821(e)(1) or 1.821(e)(2) as required by 37 CFR 1.825(b)(6)(ii); and
Statement according to item 2) a) or b) above.
Specification
The listing of references in the specification is not a proper information disclosure
statement. The specification filed on 01/23/2023 includes a list of references on pages 65-
66. 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted
for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be
incorporated into the specification but must be submitted in a separate paper." Therefore,
unless the references have been cited by the examiner on form PTO-892, they have not been
considered.
Claim Objections
Claims 1 and 10 are objected to because of the following informalities:
“An” (ln 1) should be corrected to “A”. (Claim 1, ln 1)
“one or more of a salts a reducing agent, a chelating agent, a detergent a buffering agent” should modified to “one or more of: a salt, a reducing agent, a chelating agent, a detergent, a buffering agent” (Claim 10, ln 1-2)
Appropriate correction is required.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 2-5, 9-13 and 22 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 2-3 are indefinite over the limitations “comprising MMLV-RT and Taq Pol in a 2:1 ratio” (Claim 2 ln 1) and “wherein the ratio comprises nanograms of MMLV-RT to units (U) of Taq Pol” (Claim 3 ln 1-2). It is unclear whether the ratio is indicating an activity ratio in units or a total amount ratio in nanograms, as a ratio is generally considered to of the same kind of unit, (e.g. Units of MMLV-RT to Units of Taq Pol. Furthermore, a skilled artisan would in the field expect for the prepared enzyme amounts to be described in units or units/volume (µl or ml). Claim 3 depends on claim 2.
Claims 4-5 are indefinite over the limitations “from about.”(Claim 4 ln 1 and ln 2). The phrase “from about" is vague and indefinite based on the unclear metes and bounds meant by the phrase. The phrase "from” typically indicates a minimum point. The phrase "from", however, is controverted by the term "about" which implies that values above and below are permitted. In Amgen, Inc. v. Chugai Pharmaceutical Co., 927 F.2d.1200 (CAFC 1991), the CAFC stated, "The district court held claims 4 and 6 of the patent invalid because their specific activity limitation of "at least about 160,000" was indefinite". After review, the CAFC states "We therefore affirm the district court's determination on this issue." Thus, the phrase "from about" is indefinite where the metes and bounds of the term were not defined in the specification because it is not clear if “from about” 20 ng would include amounts that were about 20 ng (e.g., 17 ng), because it is not clear if 17 ng would be from 20 ng. Furthermore, a skilled artisan would in the field expect for the prepared enzyme amounts to be described in units or units/volume (µl or ml). Claim 5 depends on claim 4.
Claim 9 recites the limitation " where the tag sequence is selected from His tag and Strep tag" in ln 1-2. There is insufficient antecedent basis for this limitation in the claim.
Claims 10-13 are indefinite over the limitations “the buffer comprises one or more of” (Claim 10 ln 1-2) and “or combinations thereof” (Claim 10 ln 3). It is unclear whether the “one or more of” is indicating at least one reagent from the whole list of reagents in claim 10 or a combination of a least one of each reagent type. Claims 11-13 depend on claim 10.
Claim 22 is indefinite over the limitations “a sequence represented by SSAG-A-L1-B-L2-C” (ln 2), “(iii) B is a protein tag as disclosed herein” (ln 4) and “(v) B is a second protein tag” (ln 5). It is unclear whether the “C” in the represented sequence is indicating the same protein tag as B, second protein tag that is different than the tag indicated in B, or the C-terminus of the sequence.
Claims 3-5 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being incomplete for omitting essential elements, such omission amounting to a gap between the elements. See MPEP § 2172.01. The omitted elements are: activity units of MMLV-RT. Enzyme reagent compositions generally describe the amount of an enzyme reagent by the measured units of activity instead of nanograms.
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 9 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. Claim 9 depends on claim 8 and claim 8 is canceled. 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.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 1-6 and 10-13 are rejected under 35 U.S.C. 101 because the claimed invention is directed to compositions that are structurally identical to the naturally occurring compositions related to natural phenomena and abstract ideas of mathematical relationships of ratios, without significantly more. The claim(s) recite(s) natural phenomena and abstract ideas. This judicial exception is not integrated into a practical application because no additional elements integrate the judicial exceptions into a practical application. The claims do not include additional elements that are sufficient to amount to significantly more than the judicial exception because no additional elements are considered significantly more than the judicial exceptions.
Claim analysis
The instant claim 1 is directed towards: An RT-PCR reagent composition comprising a wild-type Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT) and a Thermus aquaticus DNA polymerase (Taq Pol) in effective amounts and in a buffer effective for one-step RT-PCR, optionally, wherein the Tag Pol is not Platinum Tag Pol and wherein MMLV-RT and Tag Pol are in a 2:1 ratio.
The wild-type Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT) is considered a natural product related to a naturally occurring compositions of matter or synthetically created compositions that is structurally identical to the naturally occurring compositions.
The Thermus aquaticus DNA polymerase (Taq Pol) is considered a natural product related to a naturally occurring compositions of matter or synthetically created compositions that is structurally identical to the naturally occurring compositions.
Dependent claims set forth further limitations of ratios, amount of reagents and buffer reagents.
The instant claim 2 is directed towards: The composition of claim 1 comprising MMLV-RT and Taq Pol in a 2:1 ratio. The 2:1 ratio is considered an abstract idea related to a mathematical relationship.
The instant claim 3 is directed towards: The composition of claim 2, wherein the ratio comprises nanograms of MMLV-RT to units (U) of Taq Pol. The ratio is considered an abstract idea related to a mathematical relationship.
According to the 2019 Patent Eligibility Guidance an initial two step analysis is required for determining statutory eligibility.
Step 1. Is the claim directed to a process, machine, manufacture, or composition of matter? In the instant case the Step 1 requirement is satisfied as the claims are directed towards a composition of matter.
Step 2A Prong one. Does the claim recite a law of nature, a natural phenomenon or an abstract idea? Yes, natural phenomena and abstract ideas.
With regards to claim 1, the claim recites, “An RT-PCR reagent composition comprising a wild-type Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT) and a Thermus aquaticus DNA polymerase (Taq Pol) in effective amounts and in a buffer effective for one-step RT-PCR, optionally, wherein the Tag Pol is not Platinum Tag Pol and wherein MMLV-RT and Tag Pol are in a 2:1 ratio.” The wild-type Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT) and Thermus aquaticus DNA polymerase (Taq Pol) are considered natural products related to a naturally occurring compositions of matter or synthetically created compositions that is structurally identical to the naturally occurring compositions. The natural products, MMLV-RT and Taq Pol, do not possess‘ markedly different characteristics from the enzymes, MMLV-RT and Taq Pol, respectively, found in nature.
With regards to claim 2, the claim recites, “The composition of claim 1 comprising MMLV-RT and Taq Pol in a 2:1 ratio. The 2:1 ratio is considered an abstract idea related to a mathematical relationship.”
With regards to claim 3, the claim recites, “The composition of claim 2, wherein the ratio comprises nanograms of MMLV-RT to units (U) of Taq Pol. The ratio is considered an abstract idea related to a mathematical relationship.”
Step 2A prong two. Does the claim recite additional elements that integrate the judicial exception into a practical application? No, there are no additional steps that integrate the claims into a practical application.
Step 2B. Does the claim recite additional elements that are significantly more than the judicial exceptions? No, there are no additional elements that are significantly more than the judicial exceptions.
Regarding claim 1, the claim requires the combination of reagents that are well-understood, routine and conventional in the field to similar to that of Lee et al. (“Lee”; Patent App. Pub. US 20160097086 A1, April 7, 2016).
Lee discloses “…The reagent mixture comprises a ready to use reagent solution, wherein the solution comprises:… (b) a viral reverse transcriptase; and (c) at least one DNA polymerases, in a buffer suitable for use in a reverse transcription reaction, wherein the buffer comprises a co-factor metal ion and nucleoside triphosphates” (Abstract).Thus, the claim does not provide additional elements which are significantly more.
Dependent claims require amount of reagents and buffer reagents which are all routine and conventional based on Lee et al. (“Lee”; Patent App. Pub. US 20160097086 A1, April 7, 2016) in view of Lee et al. (“Lee 2014”; Patent App. Pub. EP 2354251 B1, Oct. 15, 2014), Zhao et al. (“Zhao”; Patent Pub. US 6300073 B1, Oct. 9, 2001, Lee et al. (“Lee 2018”; Patent App. Pub. US 20180312822 A1, Nov. 1, 2018). Furthermore, "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." (MPEP 2144.05).
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.
Claims 1, 4-5 and 10-11 are rejected under 35 U.S.C. 103 as being unpatentable over Lee et al. (“Lee”; Patent App. Pub. US 20160097086 A1, April 7, 2016).
Lee discloses “…The reagent mixture comprises a ready to use reagent solution, wherein the solution comprises:… (b) a viral reverse transcriptase; and (c) at least one DNA polymerases, in a buffer suitable for use in a reverse transcription reaction, wherein the buffer comprises a co-factor metal ion and nucleoside triphosphates” (Abstract).
Claim interpretations: The limitation(s) reciting “optionally” in claim 1, is interpreted as rendering the phrase thereafter as an optional limitation to the claim.
Regarding claim 1, Lee teaches a composition comprising “The reagent mixture of the present invention comprises… (b) a viral reverse transcriptase in a concentration sufficient for use in a reverse transcription reaction … wherein the viral reverse transcriptase is selected from the group consisting of… Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV RT)… and (c)one or more DNA polymerases selected from the group consisting of Thermus Aquaticus (Taq) … in a buffer suitable for use in a reverse transcription reaction” (Para. 28). “Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV RT)” reads on wildtype MMLV RT. Thus, Lee teaches a RT-PCR reagent composition comprising a wild-type Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT) and a Thermus aquaticus DNA polymerase (Taq Pol) in effective amounts and in a buffer effective for one-step RT-PCR.
Regarding claim 4-5, Lee teaches a composition wherein “2-20 units MMLV RT, 1-5 unit Taq DNA polymerase” (Para. 39). In accordance with the 112(b) rejection made above, “40 ng to about 80 ng MMLV-RT, from about 20 U to about 40 U Taq Pol, or combinations thereof” is interpreted as a composition comprising about 2-fold to 4-fold more MMLV-RT than Taq Pol. Furthermore, “about 60 ng MMLV-RT and about 30 U Taq Pol” is interpreted as a composition comprising about 2-fold more MMLV-RT than Taq Pol. Thus, Lee suggests a composition comprising from about 40 ng to about 80 ng MMLV-RT, from about 20 U to about 40 U Taq Pol, or combinations thereof; comprising about 60 ng MMLV-RT and about 30 U Taq Pol.
Regarding claim 10, Lee teaches a composition wherein “the buffer may comprise a potassium salt, a magnesium salt, nucleoside triphosphates, Dithiothreitol… non-ionic detergent” (Para. 29), “Suitable buffer compounds, such as Tris-HCl, Tris-SO.sub.4, HEPES, Tricine etc., are well known in the art.” (Para. 43), and “a chelating agent” (Para. 8). Thus, Lee teaches a composition wherein the buffer comprises one or more of a salts a reducing agent, a chelating agent, a detergent a buffering agent, a plurality of deoxynucleoside triphosphates (dNTPs), or combinations thereof.
Regarding claim 11, Lee teaches a composition wherein “K, NH4, a magnesium salt” (Para. 18), “KCl… MgCl2” (Para. 39). Thus, Lee teaches a composition wherein the one or more salts are selected from the group comprising KCl, MgCl2, and (NH4)2SO4.
Therefore, the invention as recited in claims 1, 4-5 and 10-11 are prima facie obvious over the prior art Lee et al. One of ordinary skill in the art would have had a reasonable expectation of success given the obviousness of the claim limitations in view Lee et al.. It would have been obvious to provide a RT-PCR reagent according to the limitations of the instant application claims 1, 4-5 and 10-11 based on Lee et al. (Patent App. Pub. No. US 20160097086 A1).
Claim(s) 2-3 and 12-13 are rejected under 35 U.S.C. 103 as being unpatentable over Lee et al. (“Lee”; Patent App. Pub. US 20160097086 A1, April 7, 2016) as applied to claim 1 and further in view of Lee et al. (“Lee 2014”; Patent App. Pub. EP 2354251 B1, Oct. 15, 2014).
The teachings of Lee are documented above in the rejection of claims 1, 4-5 and 10-11 under 35 U.S.C. 103. Claim 3 depends on claim 2, which depends on claim 1. Claims 12 and 13 depend on Claim 10, which depends on claim 1. Lee does not explicitly teach the limitations of claim 2-3 and 12-13.
Lee 2014 discloses The present invention is directed to compositions and methods useful for the amplification of nucleic acid molecules by reverse transcriptase-polymerase chain reaction (RT-PCR). Specifically, the invention provides compositions and methods for the amplification of nucleic acid molecules in a simplified one- or two-step RT-PCR procedure using combination of reverse transcriptase and thermostable DNA polymerase enzymes in conjunction, with sulphur-containing molecules or acetate-containing molecules (or combinations of such sulphur-containing molecules and acetate-containing molecules), and optionally bovine serum albumin. The invention thus facilitates the rapid and efficient amplification of nucleic acid molecules and the detection and quantitation of RNA molecules. The invention also is useful in the rapid production and amplification of cDNAs which may be used for as variety of industrial, medical and forensic purposes.” (Abstract)
Regarding claims 2-3, Lee 2014 teaches a composition wherein “ratio of M-MLV-RT to DNA polymerase is greater than about 3:2” (Para. 10). Lee also teaches a composition wherein “2-20 units MMLV RT, 1-5 unit Taq DNA polymerase” (Para. 39). In accordance with the 112(b) rejection made above “the ratio comprises nanograms of MMLV-RT to units (U) of Taq Pol” is interpreted as a composition wherein the ratio of MMLV-RT to Taq Pol is in units. Thus, Lee and Lee 2014 suggest a composition comprising MMLV-RT and Taq Pol in a 2:1 ratio; wherein the ratio comprises units of MMLV-RT to units (U) of Taq Pol. Lee and Lee 2014 do not explicitly teach nanograms of MMLV-RT as it is not a description of amount of enzyme in the art as discussed in the 112(b) rejection above.
Lee and Lee 2014 are both considered to be analogous to the claimed invention because they are in the same field of compositions useful for one-step RT-PCR. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the RT-PCR reagent according to the limitations of the instant application claim 1 as taught by Lee to incorporate the composition comprising a 2:1 ratio of MMLV RT to Taq pol as taught by Lee 2014 and provide a RT-PCR reagent composition in effective amounts comprising MMLV-RT and Taq Pol in a 2:1 ratio. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 2. Doing so would allow for a composition suitable for carrying out a one-step reverse transcriptase-polymerase chain (RT-PCR). (Para. 11)
Regarding claim 12, Lee 2014 teaches a composition wherein “The one or more nucleotides employed in a method of the invention may be deoxyribonucleoside triphosphates (most preferably dATP, dUTP, dTTP, dGTP or dCTP),” (Para. 16). Thus, Lee and Lee 2014 suggest a composition wherein the dNTPs are selected from the group consisting of dATP, dTTP, dGTP, and dCTP.
Regarding claim 13, Lee teaches a composition wherein “A typical kit for RT-PCR contains the following components: Reaction Buffer (20-80 mM TrisCl pH 8.5, (or 50-100 mM Tris-SO4 pH 8.9), 30-100 mM KCl. 1-2 mM MgCl.sub.2), 0.2 mM dNTPs… and stabilizer” (Para. 39). “stabilizer” reads on (NH4)2SO4. However, Lee does not explicitly teach 13.5 mM (NH4)2SO4. Lee 2014 teaches a composition comprising “18 mM (NH4)2SO4” (Para. 49; Table 1)."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." (MPEP 2144.05). Thus, Lee and Lee 2014 suggest a composition comprising 20-50 mM Tris-HCl (pH 8.5); 75-150 mM KCl; 2-4 mM MgCl2; 0.5-2 mM DTT; 200-500 µM dNTPs and 13.5 mM (NH4)2SO4.
Claim(s) 6 is rejected under 35 U.S.C. 103 as being unpatentable over Lee et al. (“Lee”; Patent App. Pub. US 20160097086 A1, April 7, 2016) as applied to claim 1 above, and further in view of over Zhao et al. (“Zhao”; Patent Pub. US 6300073 B1, Oct. 9, 2001).
The teachings of Lee are documented above in the rejection of claims 1, 4-5 and 10-11 under 35 U.S.C. 103. Claim 6 depends on claim 1. Lee does not explicitly teach the limitations of claim 6.
Zhao discloses “The field of this invention is nucleic acid amplification, and particularly one-step RT-PCR.” (Para. 2-Technichal field)
Claim interpretations: The limitation(s) reciting “and/or (c)” in claim 6, is interpreted as being limited to at least one of the limitations described as a, b or c the claim.
Regarding claim 6, Zhao teaches a composition wherein Taq Pol comprises Sequence 7, which has 99% identity to SEQ ID NO: 2 of the instant application (See attached BLAST sequence alignment). Thus, Zhao teaches a composition wherein: (a) the MMLV-RT comprises the amino acid sequence of SEQ ID NO:1 or an amino acid sequence comprising at least 85% sequence identity to SEQ ID NO:1, (b) the Taq Pol comprises the amino acid sequence of SEQ ID NO:2 or an amino acid sequence comprising at least 85% sequence identity to SEQ ID NO:2; and/or (c) the MMLV-RT and/or Taq Pol further comprise one or more tag sequences at the N-terminus and/or C-terminus.
Lee and Zhao are both considered to be analogous to the claimed invention because they are in the same field of compositions useful for one-step RT-PCR. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the RT-PCR reagent according to the limitations of the instant application claim 1 as taught by Lee to incorporate the composition comprising an amino acid sequence comprising at least 99% sequence identity to SEQ ID NO: 2 as taught by Zhao and provide a RT-PCR reagent composition wherein the Taq Pol comprises the amino acid sequence of SEQ ID NO:2 or an amino acid sequence comprising at least 85% sequence identity to SEQ ID NO:2. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 6. Doing so would allow for a composition suitable for carrying out a one-step reverse transcriptase-polymerase chain (RT-PCR).
Claim(s) 22-23 are rejected under 35 U.S.C. 103 as being unpatentable over Lee et al. (“Lee”; Patent App. Pub. US 20160097086 A1, April 7, 2016) as applied to claim 1 above, and further in view of Lee et al. (“Lee 2018”; Patent App. Pub. US 20180312822 A1, Nov. 1, 2018).
The teachings of Lee are documented above in the rejection of claims 1, 4-5 and 10-11 under 35 U.S.C. 103. Claim 22-23 depends on claim 1. Lee does not explicitly teach the limitations of claim 22-23.
Lee 2018 discloses “…are compositions, methods, and kits comprising engineered reverse transcription enzymes that exhibit several desired properties such as thermal stability, processive reverse transcription, non-templated base addition, and template switching ability…” (Abstract).
Claim interpretations: The limitation(s) reciting “optional” in claim 22, is interpreted as rendering the “L1 and L2 are optional first linkers” as an optional limitation to the claim. “SSAG” is interpreted as a flexible linker that generally comprises G/S. “SEQ ID:20” is comprised of at least one linker comprising G/S, a TEV protease cleavage sequence, a Poly-His-tag sequence and Streptavidin tag sequence which are all well-known sequences in the art.
Regarding claims 22-23, Lee 2018 suggests a composition wherein “engineered reverse transcription enzymes further comprise an affinity tag at said N-terminus or at a C-terminus of said amino acid sequence. In some instances, said affinity tag include, but are not limited to, …Strep-tag... poly(His) tag. In some instances, said affinity tag is at least 5 histidine amino acids.” (Para. 194). Lee 2018 suggests a composition wherein “In some embodiments, engineered reverse transcription enzymes further comprises a protease cleavage sequence, wherein cleavage of said protease cleavage sequence by a protease results in cleavage of said affinity tag from said engineered reverse transcription enzyme. In some instances, protease cleavage sequence is the protease cleavage sequence recognized by a protease including… tobacco etch virus (TEV)” (Para. 195). “ssag” and “any variant thereof” reads on any short flexible linker comprising mostly G/S. One of skill of the art would be motivated to include a short linker between a cleavage site and sequence of interest. Thus, Lee and Lee 2018 suggest a composition wherein the MMLV-RT sequence comprises a sequence represented by SSAG-A-L1-B-L2-C, where: (i) SSAG is an amino acid sequence or a variant thereof made by conservative substitution of the amino acids therein; (ii) A is a peptide cleave sequence, (iii) B is a protein tag as disclosed herein, (iv) L1 and L2 are optional first linkers, and (v) B is a second protein tag; and wherein the MMLV-RT comprises SSAGENLYFQGSSSHHHHHHHHGGGSAW SHPQFEK (SEQ ID NO: 20).
Lee and Lee 2018 are both considered to be analogous to the claimed invention because they are in the same field of compositions useful for one-step RT-PCR. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the RT-PCR reagent according to the limitations of the instant application claim 1 as taught by Lee to incorporate the composition comprising an amino acid sequence comprising at least 99% sequence identity to SEQ ID NO: 2 as taught by Lee 2018 and provide a RT-PCR reagent composition wherein the Taq Pol comprises the amino acid sequence of SEQ ID NO:2 or an amino acid sequence comprising at least 85% sequence identity to SEQ ID NO:2. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 22-23. Doing so would allow for preparation of MMLV-RT.
Conclusion
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure. Takahashi, M., Tehseen, M., Salunke, R., Takahashi, E., Mfarrej, S., Sobhy, M. A., Alhamlan, F. S., Hala, S., Ramos-Mandujano, G., Al-Qahtani, A. A., Alofi, F. S., Alsomali, A., Hashem, A. M., Khogeer, A., Almontashiri, N. A. M., Lee, J. M., Mon, H., Sakashita, K., Li, M., Kusakabe, T., … Hamdan, S. M. (2021). Quick and Easy Assembly of a One-Step qRT-PCR Kit for COVID-19 Diagnostics Using In-House Enzymes. ACS omega, 6(11), 7374–7386. (Whole document); pDEST8-MMLVRT-TEV-His8-Strept (Claims 22-23).
No claims are in condition for allowance.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to KENDRA R VANN-OJUEKAIYE whose telephone number is (571)270-7529. The examiner can normally be reached M-F 9:00 AM- 5:00 PM.
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/KENDRA R VANN-OJUEKAIYE/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682
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