DETAILED ACTION
The amendment filed on 11/19/2025 has been entered.
Claims 1 and 7 were amended in the claim set filed on 11/19/2025. No New matter was added.
Applicant's election with traverse of Group I (claims 1-16) in the reply filed on 05/01/2025 is acknowledged.
Claim 17 was canceled in the claim set filed on 11/19/2025. Applicant is reminded that upon the cancelation of claims to a non-elected invention, the inventorship must be corrected in compliance with 37 CFR 1.48(a) if one or more of the currently named inventors is no longer an inventor of at least one claim remaining in the application. A request to correct inventorship under 37 CFR 1.48(a) must be accompanied by an application data sheet in accordance with 37 CFR 1.76 that identifies each inventor by his or her legal name and by the processing fee required under 37 CFR 1.17(i).
Claims 1-16 in the claim set filed on 11/19/2025 are currently under examination.
Response to the Arguments
Objections to the Specification in the previously mailed non-final are maintained in light of the objection not being addressed by amendment or filing of an additional IDS.
Applicant’s arguments regarding previous rejection(s) of claim 1 under 35 U.S.C. 112 have been fully considered and are persuasive. The 35 U.S.C. 112 rejections documented in the previously mailed non-final have been withdrawn in light of applicants arguments on Pg. 5-7.
Applicant’s arguments regarding previous rejection(s) of claim(s) 1-16 under 35 U.S.C. 103 have been fully considered and are not persuasive. Applicant’s argument on Pg. 9, states that “the cited references do not teach or suggest the features of the present invention”. The 35 U.S.C. 103 rejections documented in the previously mailed non-final have been maintained and revised in light of applicants claim amendments and arguments on Pg. 7-12.
The revised rejections for claims 1-16 are documented below in this Final Office Action are necessitated by claim amendments filed on 11/19/2025.
Priority
This application is a 371 National Phase of PCT International Application No. PCT/US2020/033929, filed on May 21, 2020, which claims priority under 35 U.S.C. 119(e) to U.S. Provisional Application No. 62/850,651, filed on May 21, 2019.The priority date of claim set filed on Nov. 19, 2025, is determined to be May 21, 2019.
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 1-8 and 12-16 are rejected under 35 U.S.C. 103 as being unpatentable over Blauwkamp et al. (“Blauwkamp”, US Patent App. Pub. No. US 2018/0371538 A1, Dec. 27, 2018) in view of Chiu et al. (“Chiu”, Patent App. Pub. No. WO 2015/089496 A1, Published June 18, 2015, Filed Dec. 15, 2013), Shishkin et al. (“Shishkin”, US Patent App Pub. No. US 20160024572 A1, Jan. 28, 2016), and Tzanetakis et al. (“Tzanetakis “, (2005). The use of reverse transcriptase for efficient first-and second-strand cDNA synthesis from single-and double-stranded RNA templates. Journal of virological methods, 124(1-2), 73-77.
Blauwkamp discloses “methods of processing nucleic acids of multiple forms (single-stranded DNA, double-stranded DNA, single stranded RNA, and/or double-stranded RNA) within samples to identify the nucleic acids present within the sample”. (Para.5).
Regarding claim 1 step (a), Blauwkamp teaches a method wherein “phosphatase was added to the sample in order to remove residual phosphate groups from the 5' and 3' ends” (Para. 330). “phosphatase” is broadly interpreted as a phosphatase capable of dephosphorylating ssDNA. Thus, Blauwkamp teaches a method comprising (a) removing 5' phosphate from the nucleic acid fragment to produce a de-phosphorylated fragment.
Regarding claim 1 step (b), Blauwkamp teaches a method wherein “the single-stranded nucleic acids (DNA or RNA) may then be subjected to a first adapter ligation step 120 to append a first adapter 160 to the 3' end of the nucleic acid strand” (Para. 62) and “a single-stranded 3'-end adapter contains a phosphorylated 5'-end” (Para. 138). Blauwkamp also teaches a method wherein “phosphatase was added to the sample in order to remove residual phosphate groups from the 5' and 3' ends” (Para. 330). The P oligo is broadly interpreted as any oligonucleotide with a 5’ phosphate. “phosphatase” is broadly interpreted as a phosphatase capable of dephosphorylating ssDNA. Thus, Blauwkamp suggests a method wherein (b) ligating a P oligo DNA, a single-stranded DNA having a P oligo sequence and carrying a 5'-phosphate, to 3’-end of the de-phosphorylated ssRNA fragment to form a ssRNA-P oligo DNA strand.
PNG
media_image1.png
765
611
media_image1.png
Greyscale
Regarding claim 1 step (c), Blauwkamp teaches a method wherein “tagging … an end (a 3′ end) of the first RNA “ and “synthesizing a complementary cDNA strand from the first RNA with a reverse transcriptase” (Para. 10). Blauwkamp also teaches a method wherein “In the first replication or primer extension step (FIG. 1, 130), the enzymes used in a single reaction mixture may be able to perform the primer extension reaction against both of a DNA or RNA template with a sufficient level of replication of each. Generally, the primer extension portion of the reaction involves the addition of one or more primers (FIG. 1, 170) that recognize (hybridize to) the first adapters attached to the single-stranded DNA and/or RNA. Primer extension also involves use of a polymerase (RT, DNA polymerase)” (Para. 180; Figure 1 (see below). The P oligo is broadly interpreted as any oligonucleotide with a 5’ phosphate. The T oligo is interpreted as any oligo with a 3’ Thymine. The single-stranded DNA having a T oligo sequence that is complementary to the P oligo DNA, is interpreted as a primer, as stated in the instant claim. Thus, Blauwkamp teaches a method comprising (c) performing a first reverse transcription by using the ssRNA-P oligo DNA strand as a template and adding a T oligo DNA, a single-stranded DNA having a T oligo sequence that is complementary to the P oligo DNA, as a primer, to synthesize a complementary DNA (cDNA) strand that is complementary to the ssRNA fragment to produce a cDNA-T oligo DNA strand and thus form an initial RNA/DNA hybrid composed of said ssRNA-P oligo DNA strand and the cDNA-T oligo DNA strand.
Regarding claim 1 step (d), Blauwkamp teaches a method wherein “the adapters may be attached by a ligation reaction” (Para. 111) and “the adapter may be a RNA molecule” ( Para. 120). Blauwkamp teaches a method wherein “the first and second adapters, … can be attached using various different schemes. In some applications, the first and second adapters, or successive iterations of adapters, may be attached to ssDNA, dsDNA, ssRNA, dsRNA, DNA, RNA, or DNA/RNA hybrid molecules, or in any combination” (Para 113). Blauwkamp teaches a method wherein “the second adapter itself may behave as a primer that recognizes one or more non-templated nucleic acids” and “the second adapters may include, or may also include, one or more T residues” (Para. 179). The P oligo is broadly interpreted as any oligonucleotide with a 5’ phosphate. The T oligo is interpreted as any oligo with a 3’ Thymine. Thus, Blauwkamp teaches a method comprising (d) ligating a T oligo in an RNA form, which is a single-stranded RNA complementary to the P oligo (DNA), to 5'-end of the ssRNA-P oligo (DNA) strand in the initial RNA/DNA hybrid, to form a T oligo (RNA)-ssRNA-P oligo (DNA) strand and thus form an intermediate RNA/DNA hybrid composed of said T oligo (RNA)-ssRNA-P oligo (DNA) strand and the cDNA-T oligo (DNA) strand, having the T oligo in RNA form as a non-complementary overhang.
Regarding claim 1 step (e), Blauwkamp suggests in schematic non-templated oligo sequences at the 5' and 3 end of ssRNA and hybridized to a ss cDNA (Fig. 1, after second adapter ligation (above)) Primer extension is interpreted as reverse transcription and amplification is interpreted as transcription with a polymerase having reverse transcription activity in the Fig. 1 above. The P oligo is broadly interpreted as any oligonucleotide with a 5’ phosphate. The T oligo is interpreted as any oligo with a 3’ Thymine. Thus, Blauwkamp teaches a method comprising (e) … to obtain a complete cDNA strand having the T oligo sequence at 5'-end and the P oligo sequence at 3' end and thus form a complete RNA/DNA hybrid of said T oligo RNA-ssRNA-P oligo DNA strand and said complete cDNA strand;
However, Blauwkamp does explicitly teach all the limitations of “(e) Performing a second reverse transcription”; “using T oligo in RNA form as a non-complementary overhang as an extended template.”; “(f) removing the ssRNA fragment and the T oligo in an RNA form from the complete RNA/DNA hybrid to produce a partial, double-stranded DNA comprising said complete cDNA strand partially hybridized at its 5'-end with the P oligo DNA; and (g) performing a polymerase chain reaction (PCR) using such complete cDNA strand as a PCR template and a T oligo primer having the T oligo sequence to prime synthesis of a double-stranded DNA product (Claim 1).
Chiu discloses a method wherein TOP-PCR can amplify RNA if the RNA molecules are converted to cDNA (Pg. 11, ln 31).
Regarding claim 1 step (e), Chiu teaches a method comprising "wherein the adaptor has one 3'-T overhang and one 3'- non-A overhang" (Pg. 1 Para. 17). The adapter is interpreted as non-complementary based on T-oligo being defined as having an 3' T. Chiu also teaches that “The foregoing description of the exemplary embodiments of the invention has been presented only for the purposes of illustration and description and is not intended to be exhaustive or to limit the invention to the precise forms disclosed. Many modifications and variations are possible in light of the above teaching.” (Para. 104). Therefore, the T in the overhang is not complemented by an A in an overhang. Thus, Chiu suggests a method comprising using the T oligo in RNA form as a non-complementary overhang as an extended template.
Regarding claim 1 step (g), Chiu teaches a method comprising "performing a polymerase chain reaction (PCR) with the adaptor-ligated target nucleic acid of unknown sequence as a template and the second strand oligo with a 3'-T as a single primer to obtain a PCR product" (Pg. 2, ln 18-20). Thus, Chiu teaches a method comprising (g) performing a polymerase chain reaction (PCR) using such complete cDNA strand as a PCR template and a T oligo primer having the T oligo sequence to prime synthesis of a double- stranded DNA product.
Shishkin discloses a universal method is disclosed for the sequencing preparation of all classes of RNA (Abstract).
Regarding claim 1 step (e), Shishkin teaches a method comprising "RNA is labeled (or barcoded) by attachment, such as ligation, of a 3′ adaptor", "reverse transcription/first strand cDNA synthesis is performed with an adaptor-specific primer that specifically binds the 3′ adaptor that has been attached to the RNA", "a second adapter ligation (ssDNA/ssDNA) by 3′ linker ligation is performed. The adapter can be universal or can be barcoded", and "2nd strand synthesis can be performed, for example to make a double stranded cDNA (ds cDNA)"( Para. 59). Shishkin also teaches “Such methods and materials are illustrative only and are not intended to be limiting. Other methods and materials similar or equivalent to those described herein can be used.” (Para. 58). Thus, Shishkin teaches a method comprising (e) performing a second reverse transcription.
Tzanetakis discloses molecular characterization of eight distinct, difficult-to-clone RNA plant viruses was accomplished after the development of a reverse transcriptase-based first- and second-strand cDNA synthesis method (Abstract).
Regarding claim 1 step (f), Tzanetakis teaches a method comprising "Typical protocols for the generation of first- and second-strand cDNA call for the use of reverse transcriptase for first-strand synthesis followed by treatment with RNase H before second-strand synthesis" (Pg. 73, col. 2 para. 2). Thus, Tzanetakis teaches a method comprising (f) removing the ssRNA fragment and the T oligo in RNA form from the complete RNA/DNA hybrid to produce a partial, double-stranded DNA comprising said complete cDNA strand partially hybridized at its 5'-end with the P oligo DNA.
Blauwkamp, Chiu, Shishkin and Tzanetakis are considered to be analogous to the claimed invention because they are in the same field of measuring or testing (characterization) methods involving nucleic acids and 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 method of converting a linear, single-stranded RNA (ssRNA) fragment to a DNA fragment and amplifying the DNA fragment as taught by Blauwkamp to incorporate a method using the T oligo in RNA form as a non-complementary overhang as an extended template and the method of performing a polymerase chain reaction (PCR) using such complete cDNA strand as a PCR template and a T oligo primer having the T oligo sequence to prime synthesis of a double- stranded DNA product as taught by Chiu and provide a method to prevent adaptor self-ligation; to incorporate the method of performing a second reverse transcription as taught by Shishkin and provide a method to make a double stranded cDNA; to incorporate the method of removing the ssRNA fragment and the T oligo in RNA form from the complete RNA/DNA hybrid to produce a partial, double-stranded DNA comprising said complete cDNA strand partially hybridized at its 5'-end with the P oligo DNA as taught by Tzanetakis and provide to provide a template for efficient DNA amplification. Blauwkamp teaches that “A person skilled in the art will also recognize when new tools developed can be applied for the analysis of amplified DNA or RNA molecules. (Para. 71) The 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 of a method of converting a linear, single-stranded RNA (ssRNA) fragment to a DNA fragment and amplifying the DNA fragment according to the limitations of claim 1. Thus, the ordinary artisan would be motivated to incorporate the teachings of Chiu, Shishkin and Tzanetakis. Doing so would allow for amplification and detection of trace amounts of RNA.
The teachings of Blauwkamp, Chiu, Shishkin and Tzanetakis are documented above in the rejection of claims 1-8 and 12-16 under 35 U.S.C. 103. Claims 2-3, 6-8, 12, 14, 15 depends on claim 1. Claim 4 depends on claim 3, which depends on claim 1. Claim 13 depends on claim 12, which depends on claim 1. Claim 16 depends on claim 15, which depends on claim 1.
Regarding claim 2, Blauwkamp teaches a method wherein “the subject can be healthy. In some cases, the subject is a human patient having, suspected of having, or at risk of having, a disease or infection” (Para. 83). The sample can be from a subject who has a specific disease, condition, or infection, or is suspected of having (or at risk of having) a specific disease, condition, or infection” (Para. 88). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein the ssRNA fragment comprises a nucleic acid sequence indicative of a healthy or diseased state of a subject.
Regarding claim 3, Blauwkamp teaches a method wherein “analytes may be any type of nucleic acid including but not limited to: double-stranded (ds) nucleic acids, single stranded (ss) nucleic acids, DNA, RNA, …” (Para. 91). Blauwkamp also teaches a method wherein “nucleic acids may be derived directly from the subject, …” (Para. 91, last sent.). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein the ssRNA fragment is present in a sample from a subject.
Regarding claim 4, Blauwkamp teaches a method wherein “a sample can contain …a bodily fluid” (Para. 78, first sent.). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein the sample is obtained from a body fluid.
Regarding claim 5, Blauwkamp teaches a method wherein “a sample can contain a body fluid such as whole blood, … urine, saliva, …, respiratory secretions, vaginal fluid, anmiotic fluid, semen…” (Para. 78). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein the sample is blood, urine, saliva, tears, sweat, breast milk, nasal secretions, amniotic fluid, semen, or vaginal fluid of the subject.
Regarding claim 6, Blauwkamp teaches a method wherein “analytes may be any type of nucleic acid including but not limited to: …, cell-free RNA…,exosomes…” ( Para. 91). Exosomes are interpreted as vesicles. Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein the ssRNA fragment is cell-free RNAs (cfRNAs) or RNAs in vesicles (vc-RNAs).
Regarding claim 7, Blauwkamp teaches a method wherein the phosphorylation step occurs after 3’ ligation of adapter to RNA and/or DNA (Figure 10 -1st two seps, Phosphorylation step, after 3’ adapter ligation step). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein prior to step (d) the ssRNA-P oligo DNA strand is phosphorylated.
PNG
media_image2.png
427
313
media_image2.png
Greyscale
Regarding claim 8, Chiu teaches a method wherein T-oligo serves as the only one PCR primer” (Pg. 8, ln 6). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein in the T oligo primer is the only primer used in amplification.
Regarding claim 12, Blauwkamp teaches a method wherein “identifies the cDNA product” (Para. 160). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method further comprising detecting the amplified cDNA product.
Regarding claim 13, Blauwkamp teaches a method wherein “Non-limiting examples of detection means which can be used are various forms of sequencing, qPCR, ddPCR, microfluidic device, or microarray. (Para. 251, last sent.). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein the detecting is performed by mass spectrometry, hybridization or sequencing.
Regarding claim 14, Blauwkamp teaches a method wherein “…reaction mixture of the present disclosure has no purification steps” (Para. 240, last sent.). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach suggest a method which does not include a purification step.
Regarding claim 15 step (i), Blauwkamp teaches a method wherein some embodiments of the methods provided … is schematically illustrated in FIG. 1 (Para 61; Figure 1 (Shown above). Blauwkamp teaches a method wherein “a sample can contain …a bodily fluid” ( Para. 78, first sent.). Blauwkamp teaches a method wherein “the sample may contain … ssRNA, or any combination thereof. (Para. 61). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method for RNA assessment, comprising (i) providing a biofluid sample from a subject, wherein the biofluid includes ssRNA fragments.
Regarding claim 15 step (ii), Blauwkamp teaches a method comprising “tagging … an end ( a 3′ end) of the first RNA “ and “synthesizing a complementary cDNA strand from the first RNA with a reverse transcriptase” ( Para. 10). Blauwkamp also teaches a method wherein “In the first replication or primer extension step (FIG. 1, 130), the enzymes used in a single reaction mixture may be able to perform the primer extension reaction against both of a DNA or RNA template with a sufficient level of replication of each. Generally, the primer extension portion of the reaction involves the addition of one or more primers (FIG. 1, 170) that recognize (hybridize to) the first adapters attached to the single-stranded DNA and/or RNA. Primer extension also involves use of a polymerase (RT, DNA polymerase)” (Para. 180; Figure 1 (see 1st figure above). A single-stranded DNA having a T oligo sequence that is complementary to the P oligo (DNA), is interpreted as a primer, as stated in the instant claim. Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method, comprising (ii) performing a method of claim 1 to convert the ssRNA fragments to DNA fragments and amplify the DNA fragments;
Regarding claim 15 step (iii), Blauwkamp teaches a method wherein “Sequencing data may be used to determine genetic sequence information, ploidy states, the identity of one or more genetic variants, as well as a quantitative measure of the variants, including relative and absolute relative measures.” (Para. 281). Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method, comprising (iii) analyzing the amplified DNA fragments for measurement of one or more characteristics of the amplified DNA fragments.
Regarding claim 16, Blauwkamp teaches a method wherein “Such analytical methods including sequencing the nucleic acids as well as bioinformatics analysis of the sequencing results.” (Para. 254). Mapping and alignment are interpreted as bioinformatic analysis. Thus, Blauwkamp, Chiu, Shishkin and Tzanetakis teach a method wherein the analyzing step includes sequencing, mapping and/or alignment.
Response to Arguments
Applicant' s arguments filed 11/19/2025 (Pg. 7-12) with respect to claims 1-16 have been considered but are not persuasive. To clarify some instances argued in the response filed 11/19/2025 see responses to each argument made by Applicant below:
Applicants’ argument: “Blauwkamo does not teach or suggest using a first adapter and a second adapter having complementary sequences to each other for converting a linear ssRNA fragment to a DNA fragment and amplifying the DNA fragment… Blauwkamo's method does not achieve an RNA/DNA hybrid or a counterpart double-stranded DNA product having a homogenous (single type) adaptor with complementary sequences to each other at both termini… Chiu, Shishkin, Tzanetakis and Nai do not remedy Blauwkamo's deficiency.” (Pg. 9)
Response: In response to applicants’ argument above, Blauwkamp does teach or suggest using a first adapter and a second adapter having complementary sequences to each other for converting a linear ssRNA fragment to a DNA fragment and amplifying the DNA fragment and does achieve an RNA/DNA hybrid having a homogenous (single type) adaptor with complementary sequences to each other at both termini. Furthermore, as recited on Pg. 5-6 of the non-final office action, Blauwkamp teaches a method wherein “the single-stranded nucleic acids (DNA or RNA) may then be subjected to a first adapter ligation step 120 to append a first adapter 160 to the 3' end of the nucleic acid strand” (Para. 62) and “a single-stranded 3'-end adapter contains a phosphorylated 5'-end” (Para. 138) … Blauwkamp teaches a method wherein “tagging … an end ( a 3′ end) of the first RNA “ and “synthesizing a complementary cDNA strand from the first RNA with a reverse transcriptase” (Para. 10). Blauwkamp also teaches a method wherein “In the first replication or primer extension step (FIG. 1, 130), the enzymes used in a single reaction mixture may be able to perform the primer extension reaction against both of a DNA or RNA template with a sufficient level of replication of each. Generally, the primer extension portion of the reaction involves the addition of one or more primers (FIG. 1, 170) that recognize (hybridize to) the first adapters attached to the single-stranded DNA and/or RNA. Primer extension also involves use of a polymerase (RT, DNA polymerase)” (Para. 180; Figure 1 (see below). Blauwkamp teaches a method wherein “the adapters may be attached by a ligation reaction” (Para. 111) and “the adapter may be a RNA molecule” (Para. 120). Blauwkamp teaches a method wherein “the first and second adapters, … can be attached using various different schemes. In some applications, the first and second adapters, or successive iterations of adapters, may be attached to ssDNA, dsDNA, ssRNA, dsRNA, DNA, RNA, or DNA/RNA hybrid molecules, or in any combination” (Para 113). Blauwkamp teaches a method wherein “the second adapter itself may behave as a primer that recognizes one or more non-templated nucleic acids” and “the second adapters may include, or may also include, one or more T residues” (Para. 179).
PNG
media_image3.png
702
561
media_image3.png
Greyscale
Applicants’ argument: “Chiu fails to teach a method starting from a linear ssRNA fragment for converting ssRNA to a DNA fragment and amplifying the DNA fragment. One of ordinary skill in the art would not know how to proceed with a method starting from a linear ssRNA fragment for converting ssRNA to a DNA fragment and amplifying the DNA fragment and arrive at the present invention from Blauwkamo in combination with Chiu since no pointer is found in these cited references to do so..” (Pg. 10)
Response: 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). Furthermore, in response to applicant’s argument that there is no teaching, suggestion, or motivation to combine the references, the examiner recognizes that obviousness may be established by combining or modifying the teachings of the prior art to produce the claimed invention where there is some teaching, suggestion, or motivation to do so found either in the references themselves or in the knowledge generally available to one of ordinary skill in the art. See In re Fine, 837 F.2d 1071, 5 USPQ2d 1596 (Fed. Cir. 1988), In re Jones, 958 F.2d 347, 21 USPQ2d 1941 (Fed. Cir. 1992), and KSR International Co. v. Teleflex, Inc., 550 U.S. 398, 82 USPQ2d 1385 (2007). In this case, Blauwkamp, Chiu, Shishkin and Tzanetakis are considered to be analogous to the claimed invention because they are in the same field of measuring or testing (characterization) methods involving nucleic acids and 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 method of converting a linear, single-stranded RNA (ssRNA) fragment to a DNA fragment and amplifying the DNA fragment as taught by Blauwkamp to incorporate a method using the T oligo in RNA form as a non-complementary overhang as an extended template and the method of performing a polymerase chain reaction (PCR) using such complete cDNA strand as a PCR template and a T oligo primer having the T oligo sequence to prime synthesis of a double- stranded DNA product as taught by Chiu and provide a method to prevent adaptor self-ligation; to incorporate the method of performing a second reverse transcription as taught by Shishkin and provide a method to make a double stranded cDNA; to incorporate the method of removing the ssRNA fragment and the T oligo in RNA form from the complete RNA/DNA hybrid to produce a partial, double-stranded DNA comprising said complete cDNA strand partially hybridized at its 5'-end with the P oligo DNA as taught by Tzanetakis and provide to provide a template for efficient DNA amplification. Blauwkamp teaches that “A person skilled in the art will also recognize when new tools developed can be applied for the analysis of amplified DNA or RNA molecules. (Para. 71) The 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 of a method of converting a linear, single-stranded RNA (ssRNA) fragment to a DNA fragment and amplifying the DNA fragment according to the limitations of claim 1. Thus, the ordinary artisan would be motivated to incorporate the teachings of Chiu, Shishkin and Tzanetakis. Doing so would allow for amplification and detection of trace amounts of RNA.
Applicants’ argument: “Shishkin does not teach or suggest using a first adapter and a second adapter having complementary sequences to each other for converting a linear ssRNA fragment to a DNA fragment and amplifying the DNA fragment, and Shishkin' s method does not achieve an RNA/DNA hybrid or a counterpart double-stranded DNA product having g homogenous (single type) adaptor with complementary sequences to each other at both termini.” (Pg. 10)
Response: In response to applicants’ 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). See response above regarding the combination of the references.
Applicants’ argument: “Tzanetakis's method does not achieve an RNA/DNA hybrid or a counterpart double-stranded DNA product having a homogenous (single type) adaptor with complementary sequences to each other at both termini.” (Pg. 11)
Response: In response to applicants’ 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). See response above regarding the combination of the references.
Applicants’ argument: “The present invention is not predictable from the prior art.” (Pg. 12)
Response: In response to applicants’ argument above, as stated above in the revised rejection, the 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 of a method of converting a linear, single-stranded RNA (ssRNA) fragment to a DNA fragment and amplifying the DNA fragment according to the limitations of claim 1.
Claims 9-11 are rejected under 35 U.S.C. 103 as being unpatentable over Blauwkamp et al. (“Blauwkamp”, US Patent App. Pub. No. US 2018/0371538 A1, Dec. 27, 2018) in view of Chiu et al. (“Chiu”, Patent App. Pub. No. WO 2015/089496 A1, Published June 18, 2015, Filed Dec. 15, 2013), Shishkin et al. (“Shishkin”, US Patent App Pub. No. US 20160024572 A1, Jan. 28, 2016), and Tzanetakis et al. (“Tzanetakis “, (2005). Journal of virological methods, 124(1-2), 73-77, as applied to claim 1 above, and further in view of Nai et al. (“Nai”, (2017). Scientific reports, 7(1), 40767.
The teachings of Blauwkamp, Chiu, Shishkin and Tzanetakis are documented above in the rejection of claims 1-8 and 12-16 under 35 U.S.C. 103 rejections. Blauwkamp, Chiu, Shishkin and Tzanetakis do not explicitly teach the limitations of claims 9-11.
Nai discloses body fluid DNA sequencing is a powerful noninvasive approach for the diagnosis of genetic defects, infectious agents and diseases… we have developed T oligo-primed polymerase chain reaction (TOP-PCR) for full-length nonselective amplification of minute quantity of DNA fragments. TOP-PCR adopts homogeneous “half adaptor” (HA), generated by annealing P oligo (carrying a phosphate group at the5′ end) and T oligo (carrying a T-tail at the 3′ end), for efficient ligation to target DNA and subsequent PCR amplification primed by the T oligo alone. Using DNA samples from body fluids, we demonstrate that TOP-PCR recovers minute DNA fragments and maintains the DNA size profile, while enhancing the major molecular populations. (Abstract)
Regarding claims 9-10, Nai teaches a method wherein Table 2 indicates DNA input ranging from 10 ng- 100 ng (Table 2 col. 2, Input DNA). Nai also teaches a method wherein “(5 ng–0.01 pg) of a plasma cfDNA sample” (Figure 8 legend, Comparison of TOP-PCR to Illumina’s PCR method using serial dilutions of plasma cfDNA sample). Thus, Nai suggests a method wherein the ssRNA fragment is present as an initial input (total RNA) in an amount in a range of 0.01 ng to 100 ng or less and 0.01 ng to 10 ng or less.
Regarding claim 11, Nai teaches a method wherein Table 2 indicates DNA input ranging from 10 ng- 100 ng (Pg. 10, Table 2 col. 2, Input DNA). Nai also teaches a method wherein “(5 ng–0.01 pg) of a plasma cfDNA sample” (Pg. 11, Figure 8 legend, Comparison of TOP-PCR to Illumina’s PCR method using serial dilutions of plasma cfDNA sample). Nai suggests a method of “> 100–200 ng input gDNA” (Pg. 7 Para. 3). Thus, Nai teaches a method wherein the ssRNA fragment is present as an initial input (total RNA) in an amount in a range of 0.01 ng to 100 ng or more.
Response to Arguments
Applicant's arguments filed 11/19/2025 with respect to claims 9-11 have been fully considered but they are not persuasive. To clarify some instances argued in the response filed 11/19/2025 see responses to each argument made by Applicant below:
Applicants’ argument: “Na does not teach or suggest how to use a homogenous(single type) adaptor with complementary sequences to each other to perform a method for converting ssRNA to a DNA fragment and amplifying the DNA fragment. Na is of no help for the skilled persons to arrive at the present invention from Blauwkamo, Chiu, Shishkin and Tzanetakis.” (Pg. 11)
Response: 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). See response above regarding the combination of the references.
Conclusion of Response to Arguments
In view of the amendments, revised rejections and the above responses to arguments are documented in this Final Office Action. No claims are in condition for allowance.
Conclusion
THIS ACTION IS MADE FINAL. Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to 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.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Winston Shen can be reached at (571)272-3157. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/KENDRA R VANN-OJUEKAIYE/Examiner, Art Unit 1682 /WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682