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 .
Claims Status
Claims 6-12, 14, 15, 17, 18, 25-27, 29, 32, 33, 35, & 39 filed on 09/11/2025 are pending. The cancellation of claim 21 without prejudice to renewal in the reply filed on 09/11/2025 is acknowledged. All the amendments and arguments have been thoroughly reviewed but are deemed insufficient to place this application in condition for allowance. The following rejections are either newly applied, as necessitated by amendment, or are reiterated. They constitute the complete set being presently applied to the instant application. Response to Applicant’s argument follow. This action is FINAL.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office Action.
Any rejection not reiterated is hereby withdrawn in view of the amendments to the claims.
Claim Rejections - 35 USC § 103
Claim(s) 6, 8, 14, 15, 18, 25-27, & 32 is/are rejected under 35 U.S.C. 103 as being unpatentable over Gütig (EP 1627924 A1), as cited on the IDS dated 06/22/2022, in view of Johne (Johne et al.; Cell Press, Vol. 17, pages 205-211, April 2009), as evidenced by Longley (Longley, Bennett, & Mosbaugh; Nucleic Acids Research, Vol. 18, pages 7317-7322, November 1990).
Regarding amended claim 6, Gütig teaches a method for detecting the presence or absence cytosine methylation in DNA (status of an epigenetic modification of a target polynucleotide sequence) by chemically or enzymatically, with methylation-specific restriction enzymes, (epigenetic modification-sensitive or epigenetic modification-dependent restriction endonuclease) treating a genomic DNA sample (nucleic acid analyte) to provide a converted DNA (target polynucleotide sequence), annealing at least one methylation specific oligonucleotide carrying a non-extendable 3’ end (a single-stranded probe oligonucleotide A0) to the converted DNA in which the non-extendable 3’ terminus of the oligonucleotide is removed through pyrophosphorolysis in the case the oligonucleotide is bound to the DNA (target polynucleotide sequence) with the methylation status to be detected to create an unblocked oligonucleotide (A1) (A0 is pyrophosphorolysed in the 3’-5’ direction from the 3’ end to create at least a partially digested strand A1), then the unblocked oligonucleotide (A1) is extended and the methylation status is concluded from the presence of absence of the extended oligonucleotide product (detecting a signal derived from the products) (paragraph [0002] lines 1-2; paragraph [0012] lines 1-11; paragraph [0013] lines 1-3 &16-18; paragraph [0014] lines 1-2; paragraph [0015] lines 1-9; paragraph [0016] lines 1-4). Gütig also teaches that the detection of the methylation status (status of epigenetic modification) of the target polynucleotide sequence is performed by detecting the unblocked oligonucleotide (A1) which is ligated to a further oligonucleotide (paragraph [0023] lines 1-6). Finally, Gütig teaches the non-extendable 3’ terminus of the oligonucleotide is removed through pyrophosphorolysis in which the blocked 3’ end is removed by an enzyme that has pyrophosphorolysis activity (first reaction mixture comprises a pyrophosphorolysing enzyme) in the presence of pyrophosphate (paragraph [0018] lines 1-4).
Gütig does not teach that the partially digested strand A1 is circularized through ligation of its 3’ and 5’ ends to create A2.
Johne teaches phi29 polymerase-dependent rolling circle amplification (RCA) in which the phi29 polymerase enzyme catalyzes three degradative reactions including pyrophosphorolytic in the 3’ to 5’ direction and further that the use of phi29 polymerase RCA include the detection of probes of linear DNA molecules which after binding to target sequences are circularized by ligation (partially digested strand A1 that is partially digested by a pyrophosphorolysing enzyme (phi29 polymerase) is circularized through ligation of its 3’ and 5’ ends to create A2) (pg. 205 paragraph bridging column 1 & 2 lines 1-9; pg. 205 column 2 1st full paragraph line 17-21; pg. 206 paragraph bridging column 1 & 2 lines 1-16; pg. 206 column 2 3rd full paragraph lines 1-3; Figure 1; Figure 2). In addition, Johne teaches that the phi29 polymerase with pyrophosphorolytic activity has several features which make it most suitable for the efficient amplification of circular DNA molecules from complex biological samples (pg. 205 paragraph bridging column 1 & 2 lines 1-9).
Gütig and Johne are considered to be analogous to the claimed invention because they are all in the same field of detecting target polynucleotide sequences with pyrophosphorolysis. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of partially digesting strand A1 with a pyrophosphorolysing enzyme in Gütig to incorporate the use of a phi29 polymerase in a RCA that includes the detection of probes that are circularized by ligation (ligation of 3’ and 5’ ends) as taught in Johne because Johne teaches that doing so would provide a method to efficiently amplify circular molecules from complex biological samples.
Regarding amended claim 8, Gütig teaches a method for detecting the presence or absence cytosine methylation in DNA (status of an epigenetic modification of a target polynucleotide sequence) by chemically or enzymatically, with methylation-specific restriction enzymes, (epigenetic modification-sensitive or epigenetic modification-dependent restriction endonuclease) treating a genomic DNA sample (nucleic acid analyte) to provide a converted DNA (target polynucleotide sequence), annealing at least one methylation specific oligonucleotide carrying a non-extendable 3’ end (a single-stranded probe oligonucleotide A0) to the converted DNA in which the non-extendable 3’ terminus of the oligonucleotide is removed through pyrophosphorolysis in the case the oligonucleotide is bound to the DNA (target polynucleotide sequence) with the methylation status to be detected to create an unblocked oligonucleotide (A1), then the unblocked oligonucleotide (A1) can be ligated to a further oligonucleotide (forming A2) (A1 undergoes ligation to form A2) (first reaction mixture comprises A0, a pyrophosphorolysing enzyme, and a ligase) (paragraph [0002] lines 1-2; paragraph [0012] lines 1-11; paragraph [0013] lines 1-3 &16-18; paragraph [0014] lines 1-2; paragraph [0015] lines 1-9; paragraph [0016] lines 1-4; paragraph [0023] lines 1-6).
Gütig does not teach that the methylation-specific restriction enzymes (restriction endonucleases) and the first reaction mixture comprising A0, a pyrophosphorolysing enzyme, and a ligase are added at the same time.
However, Gütig teaches that the converted DNA (target polynucleotide sequence) is enzymatically treated with methylation-specific restriction enzymes (restriction endonucleases) and further adding a reaction mixture comprising A0, a pyrophosphorolysing enzyme, and a ligase (first reaction mixture). Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have added the restriction endonuclease and the first reaction mixture at the same time as it would be obvious to try from choosing from a finite number of identified, predictable solutions (adding the restriction nuclease and the first reaction mixture at the same time or at different times), with a reasonable expectation of success.
Regarding claim 14, Gütig teaches the epigenetic modification is cytosine methylation (paragraph [0001] lines 1-2; paragraph [0007] lines 1-10).
Regarding claim 15, Gütig teaches that cytosine methylation is detected at specific CpG positions (epigenetic modification is at CpG islands) (paragraph [0011] lines 1-4).
Regarding claim 18, Gütig teaches the detection of the methylation status (status of epigenetic modification) of the target polynucleotide sequence is performed by extending the unblocked oligonucleotide (A1) which is ligated to a further oligonucleotide (forming A2) and that this extension through linear amplification is performed by a nucleic acid polymerase in the presence of nucleoside triphosphates and further that the linear amplification requires an oligonucleotide that is complementary to the desired nucleic acid strand (products of step (b) are introduced to a second reaction mixture comprising a primer oligonucleotide that is complementary to a portion of A0, dNTPs, and an amplification enzyme) (paragraph [0022] lines 1-3; paragraph [0023] lines 1-6; paragraph [0025] lines 1-4).
Regarding claim 25, Gütig teaches that DNA polymerases can be used to activate the 3’ termini of the oligonucleotides (act as the pyrophosphorolysing enzyme) in which two or more polymerases can be used in one reaction and some preferred DNA polymerases include Tfl and Taq, in which the Taq DNA polymerase has been shown to have 5’ to 3’ exonuclease activity, as evidenced by Longley (abstract of Longley lines 1-2; pg. 7317 of Longley column 2 1st full paragraph lines 1-10), (first reaction mixture further comprises a 5’-3’ exonuclease) (paragraph [0019] lines 1-2 & 9-10).
Longley teaches that the Taq DNA polymerase has been shown to have 5’ to 3’ exonuclease activity (abstract lines 1-2; pg. 7317 column 2 1st full paragraph lines 1-10).
Regarding claim 26, Gütig teaches the oligonucleotide with a non-extendable 3’ terminus that is bound to the converted DNA (target polynucleotide sequence) (A1) is treated with an enzyme that has pyrophosphorolysis activity in the presence of pyrophosphate (a specific form of a phosphatase) (paragraph [0018] lines 1-4; paragraph [0020] lines 1-2).
Regarding claim 27, Gütig teaches the oligonucleotide with a non-extendable 3’ terminus that is bound to the converted DNA (target polynucleotide sequence) (A1) is treated with an enzyme that has pyrophosphorolysis activity in the presence of pyrophosphate and can also be treated with a 3’ exonuclease (products of step (b) are treated with a pyrophosphatase or exonuclease) (paragraph [0018] lines 1-4; paragraph [0020] lines 1-2).
Regarding amended claim 32, Gütig teaches that DNA polymerases with pyrophosphorolysis activity can be used to activate the 3’ termini of the oligonucleotides and that the detection of the methylation status (status of epigenetic modification) of the target polynucleotide sequence is performed by extending the unblocked oligonucleotide (A1) which is ligated to a further oligonucleotide and that this extension through amplification is performed by a nucleic acid polymerase (paragraph [0019] lines 1-3; paragraph [0022] lines 1-3).
Johne teaches phi29 polymerase-dependent rolling circle amplification (RCA) in which the phi29 polymerase enzyme catalyzes three degradative reactions including pyrophosphorolytic in the 3’ to 5’ direction and further that the use of phi29 polymerase RCA include the detection of probes of linear DNA molecules which after binding to target sequences are circularized by ligation (pyrophosphorolysing enzyme (phi29 polymerase) pyrophosphorolyses in the 3’-5’ direction and amplifies A2) (pg. 205 paragraph bridging column 1 & 2 lines 1-9; pg. 205 column 2 1st full paragraph line 17-21; pg. 206 paragraph bridging column 1 & 2 lines 1-16; pg. 206 column 2 3rd full paragraph lines 1-3; Figure 1; Figure 2).
Claim(s) 7, 9, & 39 is/are rejected under 35 U.S.C. 103 as being unpatentable over Gütig (EP 1627924 A1), as cited on the IDS dated 06/22/2022, and Johne (Johne et al.; Cell Press, Vol. 17, pages 205-211, April 2009), as applied to claims 6, 8, 14, 15, 18, 25-27, & 32 above, and further in view of Cohen-Karni (Cohen-Karni et al.; PNAS, Vol. 108, pages 11040-11045, May 2011).
The teachings of Gütig and Johne with respect to claim 6 are discussed above.
Regarding amended claim 7, Gütig and Johne does not teach that the enzymatic treatment, with methylation-specific restriction enzymes (restriction endonuclease), cleaves the target polynucleotide sequence in which a target epigenetic state is present.
Cohen-Karni teaches that MspJI is a modification-dependent restriction enzyme that recognizes and cleaves at a fixed distance away from specific cytosine C5 modification (CpG) site (methylation or hydroxymethylation) (restriction endonuclease cleaves target polynucleotide sequence in which the epigenetic state is present) (abstract lines 1-21). Cohen-Karni also teaches that MspJI and its family of enzymes provides powerful tools for direct interrogation of the epigenome (abstract lines 1-21).
Gütig, Johne, and Cohen-Karni are considered to be analogous to the claimed invention because they are all in the same field of detection of epigenetic modification through methylation-specific enzyme treatment of the sample. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of treating a genomic DNA sample with a methylation-specific restriction enzyme in Gütig to incorporate the use of the specific modification-dependent restriction enzyme of MspJI that cleaves the target polynucleotide sequence when the modification is present as taught in Cohen-Karni because Cohen-Karni teaches that doing so would provide a powerful tool for direct interrogation of the epigenome.
Regarding amended claim 9, Gütig teaches that the methylation specific oligonucleotide carries a non-extendable 3’ end (A0) (chemical modification at or close to its 3’ end) and that the methylation specific oligonucleotide carrying a non-extendable 3’ end (A0) only becomes unblocked, where the non-extendable 3’ terminus is removed, in the case that the oligonucleotide is bound to the converted DNA (target polynucleotide sequence) (modification is removed through annealing of A0 with restriction endonuclease treated converted DNA) (paragraph [0012] lines 1-11; paragraph [0014] lines 1-2; paragraph [0015] lines 1-9; paragraph [0016] lines 1-4; paragraph [0023] lines 1-6).
Gütig and Johne does not teach that the enzymatic treatment, with methylation-specific restriction enzymes (restriction endonuclease), cleaves the modification or mismatch of A0.
Cohen-Karni teaches that MspJI is a modification-dependent restriction enzyme that recognizes and cleaves at a fixed distance away from specific cytosine C5 modification (CpG) site (methylation or hydroxymethylation) in which MspJI recognizes 5-methylcytosine or 5-hydroxymethylcytosine and cleaves the sequence at specific positions on the 3’ side of the modified base (the 3’ end modification or mismatch is removed through cleavage of A0 by the restriction endonuclease) (abstract lines 1-21; pg. 11041 column 1 3rd full paragraph lines 1-4).
Gütig, Johne, and Cohen-Karni are considered to be analogous to the claimed invention because they are all in the same field of detection of epigenetic modification through methylation-specific enzyme treatment of the sample. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of treating a genomic DNA sample with a methylation-specific restriction enzyme in Gütig to incorporate the use of the specific modification-dependent restriction enzyme of MspJI that cleaves the target polynucleotide sequence at specific positions of the 3’ side of the modified base when the modification is present as taught in Cohen-Karni because Cohen-Karni teaches that doing so would provide a powerful tool for direct interrogation of the epigenome.
Regarding amended claim 39, Gütig and Johne does not teach that the restriction endonuclease is MspJI or LpnPI.
Cohen-Karni teaches that MspJI is a modification-dependent restriction enzyme that recognizes and cleaves at a fixed distance away from specific cytosine C5 modification (CpG) site (methylation or hydroxymethylation) (abstract lines 1-21). Cohen-Karni also teaches that MspJI and its family of enzymes provides powerful tools for direct interrogation of the epigenome (abstract lines 1-21).
Gütig, Johne, and Cohen-Karni are considered to be analogous to the claimed invention because they are all in the same field of detection of epigenetic modification through methylation-specific enzyme treatment of the sample. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of treating a genomic DNA sample with a methylation-specific restriction enzyme in Gütig to incorporate the use of the specific modification-dependent restriction enzyme of MspJI as taught in Cohen-Karni because Cohen-Karni teaches that doing so would provide a powerful tool for direct interrogation of the epigenome.
Claim(s) 10, 11, & 33 is/are rejected under 35 U.S.C. 103 as being unpatentable over Gütig (EP 1627924 A1), as cited on the IDS dated 06/22/2022, and Johne (Johne et al.; Cell Press, Vol. 17, pages 205-211, April 2009), as applied to claims 6, 8, 14, 15, 18, 25-27, & 32 above, and further in view of Tost (Tost; DNA Methyltransferases - Role and Function Chapter, pages 343-430, November 2016).
The teachings of Gütig and Johne with respect to claim 6 are discussed above.
Regarding claim 10, Gütig teaches that DNA can be chemically or enzymatically treated in which the enzymatic treatment comprises treatment with methylation-specific restriction enzymes and then amplified and analyzed in different ways (paragraph [0005] lines 1-2).
Gütig and Johne does not teach that the target polynucleotide sequence is selectively amplified in (a) with MS-MLPA.
Tost teaches that MS-MLPA requires two oligonucleotides with universal primer binding sites that are annealed to a target region (target polynucleotide sequence) and ligated in the case of target complementary and that a methylation-specific enzyme (restriction endonuclease) is added to the ligation reaction to digest the amount ligated product and does not rely on bisulfite conversion (pg. 380 1st full paragraph lines 1-14).
Gütig, Johne, and Tost are considered to be analogous to the claimed invention because they are all in the same field of amplification techniques with methylation-specific enzyme treatment of the sample. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of enzymatically treating DNA in Gütig to incorporate the further selectively amplifying the target polynucleotide sequence with MS-MLPA with the enzymatic treatment of the DNA as taught in Tost because Tost teaches that doing so would provide a method to amplify the target (target polynucleotide sequence) without relying on bisulfite conversion.
Regarding claim 11, Gütig teaches that DNA can be chemically or enzymatically treated in which the enzymatic treatment comprises treatment with methylation-specific restriction enzymes and then amplified and analyzed in different ways (paragraph [0005] lines 1-2).
Gütig and Johne does not teach that the products of (a) undergo PCR prior to (b).
Tost teaches that PCR amplification can be performed following methylation-specific restriction digestion (restriction endonuclease) (products of (a) undergo PCR prior to (b)) and that this method requires less DNA and no-prior bisulfite treatment and provides a rapid screening tool for differential methylation (pg. 380-381 paragraph bridging pg. 380 & pg. 381 lines 1-12).
Gütig, Johne, and Tost are considered to be analogous to the claimed invention because they are all in the same field of amplification techniques with methylation-specific enzyme treatment of the sample. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of enzymatically treating DNA in Gütig to incorporate the PCR amplification of the target polynucleotide sequence after treatment with the methylation-specific restriction enzyme as taught in Tost because Tost teaches that doing so would provide a method to amplify the target (target polynucleotide sequence) without relying on bisulfite conversion and providing a rapid screening tool for measuring differential methylation.
Regarding claim 33, Gütig teaches the detection of the methylation status (status of epigenetic modification) of the target polynucleotide sequence is performed by extending the unblocked oligonucleotide (A1) which is ligated to a further oligonucleotide (forming A2) and that this extension through linear amplification can be detected by incorporation of a labelled nucleotide (probe) or by detecting the binding or incorporation of a dye or spectral material (fluorescent binding dyes) (paragraph [0022] lines 1-3; paragraph [0023] lines 1-6; paragraph [0025] lines 1-4).
Gütig and Johne does not teach that the detection is achieved using one or more oligonucleotide fluorescent binding dyes or molecular probes in which an increase in signal over time resulting from the generation of amplicons of A2 is used to infer the concentration of the target polynucleotide sequence.
Tost teaches that quantification of targets (target polynucleotide sequence) can be detected following amplification through monitoring an increase in fluorescence with intercalating dyes as a method to monitor the target sequence in real-time (an increase in signal over time is used to infer the concentration of the target polynucleotide sequence) (pg. 380-381 paragraph bridging pg. 380 & pg. 381 lines 1-12).
Gütig, Johne, and Tost are considered to be analogous to the claimed invention because they are all in the same field of amplification techniques with methylation-specific enzyme treatment of the sample. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of extending the unblocked oligonucleotide (A1) which is ligated to a further oligonucleotide (forming A2) in which this extension through linear amplification can be detected by incorporation of a dye in Gütig to incorporate monitoring an increase in signal of the fluorescence with intercalating dyes as taught in Tost because Tost teaches that doing so would provide a method to monitor the target sequence in real-time.
Claim(s) 12 & 17 is/are rejected under 35 U.S.C. 103 as being unpatentable over Gütig (EP 1627924 A1), as cited on the IDS dated 06/22/2022, and Johne (Johne et al.; Cell Press, Vol. 17, pages 205-211, April 2009), as applied to claims 6, 8, 14, 15, 18, 25-27, & 32 above, and further in view of Rauch (Rauch & Pfeifer; Handbook of Epigenetics The New Molecular and Medical Genetics, Chapter 9, pages 135-147, 2011).
The teachings of Gütig and Johne with respect to claim 6 are discussed above.
Regarding amended claims 12 & 17, Gütig and Johne does not teach prior to step (a) enriching for the nucleic acid analyte (see claim 12) with one or more methyl-binding proteins (see claim 17).
Rauche teaches a method for purifying either restriction-enzyme cut or sonicated genomic DNA with the high affinity MBD2b/MBD3L1 complex (methyl-binding proteins) that specifically binds to the methylated genomic fragments (enriching for the nucleic acid analyte with one or more methyl-binding proteins) (pg. 143 1st full paragraph lines 1-15). Rauche also teaches that is easy to then purify the MBD2b/MBD3L1 complex containing methylated DNA with magnetic beads (pg. 143 1st full paragraph lines 1-15).
Gütig, Johne, and Rauche are considered to be analogous to the claimed invention because they are all in the same field of restriction enzyme treatment of the methylated target sequence in a sample. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of detecting cytosine methylation (epigenetic modification) in Gütig to incorporate the enrichment of the epigenetically modified or unmodified target polynucleotide sequence with a MBD2b/MBD3L1 complex as taught in Rauche because Rauche teaches that doing so would provide a high affinity and easy method to purify restriction-enzyme cut methylated DNA (target polynucleotide sequence).
Claim(s) 29 is/are rejected under 35 U.S.C. 103 as being unpatentable over Gütig (EP 1627924 A1), as cited on the IDS dated 06/22/2022, and Johne (Johne et al.; Cell Press, Vol. 17, pages 205-211, April 2009), as applied to claims 6, 8, 14, 15, 18, 25-27, & 32 above, and further in view of Raine (Raine et al.; Nucleic Acids Research, Vol. 45, pages 1-15, October 2016).
The teachings of Gütig and Johne with respect to claims 6 & 18 are discussed above.
Regarding amended claim 29, Gütig and Johne does not teach splint oligonucleotide D.
Raine teaches a splinted ligation adapter tagging method for generating a library for assessing genome wide methylation profiles in which chemically treated methylated sequences have an adapter ligated to the 3’ end of the target methylated sequence (A1) and further a splint oligonucleotide (oligonucleotide D) is annealed to the adapter and the 3’ termini of the target methylated sequence (A1) (splint oligonucleotide D is complementary to 3’ end of A1 and a portion of the adapter that A1 is ligated to) in which there is a modification at the 3’ termini of the splint oligonucleotide (oligonucleotide D) (abstract lines 13-21; Figure 1 right most column). In addition, Raine teaches that this method provides a straightforward and cost efficient approach for assessing methylation status (pg. 13 column 2 2nd full paragraph lines 8-9).
Gütig, Johne, and Raine are considered to be analogous to the claimed invention because they are all in the same field of assessing the status epigenetically modified sequences. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of detecting cytosine methylation (epigenetic modification) in Gütig to incorporate the use of a splint oligonucleotide (oligonucleotide D) that is annealed to the adapter and the 3’ termini of the target methylated sequence (A1) as taught in Raine because Raine teaches that doing so would provide a straightforward and cost effective method for assessing methylation status.
Claim(s) 35 is/are rejected under 35 U.S.C. 103 as being unpatentable over Gütig (EP 1627924 A1), as cited on the IDS dated 06/22/2022, and Johne (Johne et al.; Cell Press, Vol. 17, pages 205-211, April 2009), as applied to claims 6, 8, 14, 15, 18, 25-27, & 32 above, and further in view of Ding (U.S. Patent Application Publication US 2019/0271035 A1), as cited in the IDS dated 08/31/2022.
The teachings of Gütig and Johne with respect to claim 6 are discussed above.
Regarding amended claim 35, Gütig and Johne does not teach multiple probes A0 are employed each selective for a different target polynucleotide sequence.
Ding teaches a method for multiplex pyrophosphorolysis that can amplify multiple targets (different target polynucleotide sequences) in a single reaction through the use of a plurality of pairs of forward and reverse blocked primers for pyrophosphorolysis (the plurality of target polynucleotides has an identification region that the plurality of forward and reverse blocked primers identify) and then amplifying the multiple targets in one reaction (detection of one or more of the identification regions) (abstract lines 1-6; paragraph [0019] lines 1-5; paragraph [0021] lines 1-17).
Gütig, Johne, and Ding are considered to be analogous to the claimed invention because they are all in the same field of detecting target polynucleotide sequences with pyrophosphorolysis. Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of detecting a single target polynucleotide sequence through the detection of the extension of A2 in Gütig and Johne to incorporate the detection of multiple different target polynucleotide sequences with plurality of pairs of forward and reverse blocked primers for pyrophosphorolysis as taught in Ding because Ding teaches that doing so would provide a method to detect a plurality of target polynucleotide sequences in a single reaction.
Response to Arguments
The response traverses the rejection. The response asserts that Gütig fails to disclose the element of A1 circularizing through ligation of its 3’ and 5’ ends and, that as Gütig is deficient in this element, that the other references cited in the rejections fail to meet Gütig’s deficiencies and, as such, a prima facie case of obviousness has not been established. This argument has been thoroughly reviewed but was not found persuasive as the prior art of Gütig and Johne, as applied to newly amended claim 6 as necessitated by amendment, appreciates the use of a phi29 polymerase, a pyrophosphorolytic enzyme, in a RCA that includes the detection of probes that are circularized by ligation (partially digested strand A1 that is partially digested by a pyrophosphorolysing enzyme (phi29 polymerase) is circularized through ligation of its 3’ and 5’ ends to create A2).
The response also asserts that even if a prima facie case of obviousness had been established that Gütig disparages enzyme-based methods in paragraph 10 and, as such, one of ordinary skill in the art would not be led to the present method, which uses exactly the types of enzymes that Gütig seeks to avoid, by the prior art. This argument has been thoroughly reviewed but was not found persuasive as paragraph [0010] of Gütig teaches that the method described uses methylation specific restriction enzymes, the same type of enzymes used in the instant method, and therefore Gütig does not seek to avoid the use of epigenetic modification-sensitive or epigenetic modification-dependent restriction endonucleases as currently claimed.
For these reasons, and the reasons already made of record and modified to address the claims as currently amended, the rejections are maintained and applied to the newly amended claims.
Conclusion
Claims 6-12, 14, 15, 17, 18, 25-27, 29, 32, 33, 35, & 39 are rejected.
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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/BAILEY BUCHANAN/Examiner, Art Unit 1682
/JEHANNE S SITTON/Primary Examiner, Art Unit 1682