DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Status of Claims
Applicant's amendment filed 06/29/2026 is acknowledged. Claims 13-17 have been amended. Claims 1-12 remain withdrawn. Claims 1-17 are pending in the instant application and claims 13-17 are the subject of this final office action.
All of the amendments and arguments have been reviewed and considered. Any rejections or objections not reiterated herein have been withdrawn in light of amendments to the claims or as discussed in this office action.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Previous Rejection
Status of Prior Rejections/Objections:
The 112(b) rejections to claim(s) 14 and 16-17 is/are withdrawn in view of the amendments to the claims. See new 112(b) rejections and the objection.
The prior art rejection(s) under 35 USC 102 directed to claim(s) 13-16 as being anticipated by Giannoukous are withdrawn in view of the amendments.
The prior art rejection(s) under 35 USC 102 directed to claim(s) 13-16 as being anticipated by Steemers are withdrawn in view of the amendments.
The prior art rejection(s) under 35 USC 103 directed to claim(s) 15-17 as being unpatentable over Steemers, Giannoukous, and Shokralla are withdrawn in view of the claim amendments. See new art rejections.
New Ground(s) of Rejections
The new ground(s) of rejections were necessitated by applicant’s amendment of the claims.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Claim Objections
Claims 14 and 16 are objected to because of the following informalities:
Claims 14 and 16 recite “Seq-1”. This term creates confusion with SEQ ID NOs. Applicant may consider “a sequencing primer annealing sequence” (e.g., pg. 14, para 2).
Claim 14 recites “the Seq-1 is” in line 4 of the claim ; this should be “the Seq-1 primer is”.
Appropriate correction is required.
Claim Interpretation
In evaluating the patentability of the claims presented in this application, claim terms have been given their broadest reasonable interpretation (BRI) consistent with the specification, as understood by one of ordinary skill in the art, as outlined in MPEP 2111.
Regarding claims 13-17, claims 13 and 15 recite “wherein the amplification primer sequence comprises a reverse transcriptase (RT) primer adapted for reverse transcription of an RNA sequence into a cDNA sequence by reverse transcriptase” and “wherein the amplification primer sequence is adapted for both DNA and RNA co-amplification”. No definition of specific structures required for “reverse transcriptase” primers or “amplification primer sequences” DNA and RNA co-amplification was identified in the disclosure, nor do the facts of record identify specific structures that limit such amplification primer sequences under the broadest reasonable interpretation. Exemplary RT primers may be found on pg. 27, para 2.
As such, the amplification primer sequences have been given the broadest reasonable interpretation according to MPEP 2111.04 and the disclosure.
Claim Rejections - 35 USC § 112(b)
Claim 13-17 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Regarding claim 13, the amended claim recites “A set of oligonucleotide adapters …, wherein each adapter comprises an amplification sequence, a DNA-specific or RNA-specific barcode, a … UMI… sequence, and an annealing sequence, wherein one oligonucleotide adapter has DNA-specific barcode and the other oligonucleotide adapter has an RNA-specific barcode, wherein the amplification primer sequence comprises a … RT…primer…wherein the RT primer comprises a priming sequence, the RNA-specific barcode, and a 6-nucleotide UMI…”.
In the first wherein clause (“wherein each adapter … annealing sequence”), the amplification sequence and barcode recited as separate elements. In the newly amended wherein clauses beginning with “wherein the amplification primer sequence comprises a … RT…primer”, the RT primer comprised in amplification primer is recited as comprising the RNA-specific barcode, and a 6-nucleotide UMI. There is insufficient antecedent basis for this limitation in the claim.
First, as the RNA-specific barcode (line 8) is recited as “the” RNA-specific barcode, it is not clear whether this is intended as a second RNA-specific barcode of the same sequence or is the previously recited RNA-specific barcode as the RNA-specific barcode of line 3.
Second, given the presence of the RNA-specific barcode, it is not clear whether this RT primer is intended to apply only to “the other oligonucleotide adapter [with] an RNA-specific barcode” or also the “one oligonucleotide adapter [with] DNA-specific barcode”.
Third, it is not clear whether the “a 6-nucleotide UMI” is intended to be the same “a … UMI…sequence” as previously recited (line 3) or a second UMI.
Claim 14 is indefinite for depending from claim 13 and not rectifying the deficiency.
Regarding claim 15, as in claim 13, it is not clear whether the RNA-specific barcode of the RT-primer is the same as the RNA-specific barcode of line 3 or a second instance, wherein it is not clear how the artisan would choose between a DNA-specific or RNA-specific if the former.
Second, as above, it is not clear whether the UMIs are intended to be the same or different.
These are, at least in part, antecedent basis rejections.
Claims 16-17 are indefinite for depending from claim 15 and not rectifying the deficiency.
Claim Rejections - 35 USC § 101
Claims 13-16 are rejected under 35 U.S.C. 101 because the claimed invention is directed to judicial exception(s) without significantly more. The claim(s) recite(s) natural products. This judicial exception is not integrated into a practical application. The claim(s) does/do not include additional elements that are sufficient to amount to significantly more than the judicial exception.
The following three inquiries are used to determine whether a claim is drawn to patent-eligible subject matter:
Step 1. Is the claim directed to a process, machine, manufacture, or composition of matter?
Yes, the claims are directed to a composition of matter.
Step 2A, prong 1. Does the claim recite a law of nature, a natural phenomenon, or an abstract idea (recognized judicial exceptions)?
The claims recite a natural product. The structures required by claims 13 and 15 are “an amplification primer sequence”, “a DNA-specific or RNA-specific barcode”, “a unique molecular identifier sequence”, and “an annealing sequence”.
As amended, the claims now require the amplification primer sequence comprises a “reverse transcriptase … primer adapted for reverse transcription of an RNA sequence into cDNA by a reverse transcriptase” and that the “amplification primer sequence is adapted for both DNA and RNA co-amplification”. See interpretation above.
Under the best possible interpretation of the “wherein the RT primer” clause, the claim further requires that the UMI region be 6 nucleotides long.
Claim 13 requires that one oligonucleotide of a set have one “DNA-specific” barcode and another have an “RNA-specific” barcode.
Claim 14 recites that the adapters further comprise “a mosaic and a Seq-1 primer”. Claim 16 recites that the adapter further comprises “a mosaic and/or a Seq-1 primer”. As discussed, in the 112(b), these have been interpreted to be species of the annealing sequence and the amplification primer sequence, respectively.
An amplification primer sequence, including one adapted for reverse transcription, may be a random hexamer primer sequence. A “DNA-specific” barcode may be a SNP and the “RNA-specific” barcode may be the other. An UMI may be a de novo-acquired SNP within a 6 n-nucleotide region. The annealing sequence may be a downstream or upstream segment of the nucleic acid, wherein the function of annealing is met by annealing to the complementary strand.
Where the mosaic (sequence) is amended to be defined to be a recognition sequence for a Tn5 transposase in claims 14 and 16, the specification provides the exemplary Tn5 recognition sequence SEQ ID NO: 8 (pg. 14, para 2). This limitation thus may comprise, for example, regions of a genome with naturally occurring sequences of such Tn5 recognition sequences, e.g., a region of the P. aeruginosa genome (as aligned using NCBI BLAST):
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The Seq-1 annealing sequence is not further defined by the specification and may comprise a further sequence segment of the sequence (e.g., overlapping in the opposite direction or in the same direction).
Even under the alternative interpretation that further requires a second instance of the RNA barcode and a (second) UMI that is 6-nt, as the RNA barcode may be, for example, any nucleotide and a second UMI may be any other de novo-acquired SNP within a region of adequate length, such would not be markedly different than sequence expected to be found naturally occurring in the genome of P. aeruginosa or others that contain such a Tn5 recognition sequence. Where primer and annealing sequences or regions are claimed, it is noted that such functional limitations have been interpreted broadly (e.g., random hexamer sequences which may encompass any sequence of such a number of bases). It is further noted that the claims recite “comprises” and thus such naturally occurring regions may encompass additional elements/sequence.
None of the structures claimed require non-natural sequences/structures or features that would have markedly different characteristics from such natural sequences.
The courts have found that, even should the product be directed to single-stranded oligonucleotides, such would not be a markedly different characteristic. See MPEP 2106.04(c)(II). As such, they encompass such products of nature.
Step 2A, prong 2. Is the judicial exception(s) integrated into a practical application?
No. The claims are limited to the set of oligonucleotide adapters and under the best possible interpretation a sequence thereof or a primer sequence may overlap a portion thereof. There is no requirement that they have additional components or sequences other than what may be found in nature.
Step 2B. Does the claim amount to significantly more?
No. As above, the claims encompass only structures that may encompass what may be found in nature. As such, the claims as a whole do not encompass “significantly more” than the natural product. Nor are the claimed structures markedly different than the natural product.
Claim Rejections - 35 USC § 103
Claim(s) 13016 is/are rejected under 35 U.S.C. 103 as being unpatentable over Steemers (WO 2020/112604 A2; published 06/04/2020; priority to 11/30/2018).
Regarding claims 13 and 15, Steemers teaches a set of adapters compatible with use in a “co-assay” (Fig. 4; see para [0015]) comprising: 5’-A14-barcode-UMI-(T)n-3’
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Steemers teaches that the set of adapters may have different barcodes for identifying different analytes (para [0055]), i.e., “RNA-specific” and “DNA-specific” barcodes.
Steemers teaches that a barcode can have any length from 1 to greater than 50 including 6 nucleotides and choosing a desired length sufficient to be unique within a plurality of barcodes in a population and/or within a population and target nucleic acids being interrogated (para [0056]). Steemers teaches an “index” includes a sequence of nucleotides that can be used as a molecular identifier and/or barcode to tag a nucleic acid and/or to identify the source of the nucleic acid (para [00119]).
Steemers teaches combinatorial indexing to add additional “tags” [e.g., annealing sequences and/or amplification primer sequences; see para (0126) and claim 12] and barcodes (Fig. 11; para [0022-23], [0092], [0101]). Steemers teaches that combinatorial indexing allows for resolution with increased throughput and that the index can be used for sample identification or experimental conditions (para [0092]).
Regarding claims 14 and 16, Steemers teaches that adaptors may comprise a primer sequence, an anchor sequence [i.e., annealing sequence], a universal sequence [i.e., additional primer sequence, e.g., a mosaic and a Seq-1 primer], an index sequence (e.g., UMI), and a barcode sequence (para [0050]).
Steemers teaches an A14-ME sequence (para [0052]) and that it is useful as a bidirectional primer binding site for a Read1 primer and Index1 primer in Fig. 12:
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See also instant pg. 14, para 2 regarding Read 1 comprising mosaic and Seq-1.
Steemers teaches that the ME sequence is a transposon end sequence (para [0049]) and that the transposase may be Tn5 (para [0048]); see SEQ ID NO: 3, which teaches ME’, which is identical to the Tn5 recognition sequence instant SEQ ID NO: 8 (instant pg. 14, para 2).
Steemers teaches a nucleic acid library where the first and second tags are the same (claim 10) and the second tag comprises a transposase-specific element (claim 13).
Therefore, in the adapter(s) of Steemers, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have utilized a 6-nucleotide UMI in an adapter or set of adapters (instant claims 13 and 15), as such is taught by Steemers as a routinely optimizable variable within range for barcode such wherein the artisan chooses a desired length sufficient to be unique given a chosen population of molecules, wherein Steemers further teaches that UMIs and barcodes are used for the same purpose of indexing and such rationales for optimization would be obvious to apply to molecular identifiers as well as barcodes.
In so far as the claims may be interpreted to require the adapter or set of adapters to comprise a second analyte index and second UMI within the amplification primer sequence, the amplification primer seq of Fig. 4 of Steemers by be considered to be the region encompassing the barcode and UMI in Fig. 4 (instant claims 13 and 15), wherein it further would be obvious to the POSITA before the EFD of the claimed invention to have appended additional indices as claimed, motivated by the teachings of Steemers of using such (e.g., with intervening primer/annealing tag sequences) for combinatorial indexing for sample identification or experimental conditions, and because such further represents a duplication of parts, as discussed in MPEP 2144.04(VI)(B). In so far as the combinatorial indexing of Steemers may represent an addition of separate tag + barcode pieces, the assembly of such adapters represents only the making integral and thus would be obvious to the POSITA before the EFD of the claimed invention for at least that reason, as described in MPEP 2144.04(V)(B).
It also would have been obvious to the POSITA before the EFD of the claimed invention to have substituted the annealing sequence (e.g., A14 of Fig. 4) for a tag/annealing sequence comprising a transposon-specific sequence (e.g., ME’), as taught by Steemers, and further for that to be A14ME or A14’ME’, motivated by the teachings of Steemers that it is useful as a known bidirectional primer annealing site. It is noted that upon such substitution, the A14 or A14’ may be considered a “Seq-1” primer/annealing sequence, the ME or ME’ may be considered the Tn5 recognition mosaic region and the entire region (which is taught as two separate primer annealing regions) may be considered an annealing region in either direction (instant claims 14 and 16).
There would have been a strong expectation of success as each of the elements is taught by Steemers and such would have been predictable as routine design choices in combining these known elements according to established functions with no unexpected outcomes.
Claim(s) 17 is/are rejected under 35 U.S.C. 103 as being unpatentable over Steemers (WO 2020/112604 A2; published 06/04/2020; priority to 11/30/2018) as applied to claims 13-16 above, and further in view of EURx (EURx. Anchored Oligo(dT)20 VN Primer. Gdańsk, Poland: EURx Ltd.; 2019 [cited 2026 Sept 1]. Available from: https://eurx.com.pl/docs/specs/en/e0106.pdf) and Tang (Tang, E., et al. Development of High Throughput Processes for Constructing Illumina Libraries. Lawrence Berkeley National Laboratory. LBNL Report#: LBNL-3270E Poster. Retrieved from https://escholarship.org/uc/item/3gw4k7tk).
Regarding claim 17, Steemers further teaches the sequence B15 SEQ IQ NO: 5 is GTCTCGTGGGCTCGG (para [0052]) and utilizing it as an adaptor ending sequence on the other adapter used with the one comprising A14 in Fig. 4.
Steemers teaches that the barcodes may alternatively be chosen to be 5 nucleotides (para [0056]).
Steemers does not explicitly teach that the adapter comprises:
SEQ ID NO: 1, i.e., 5’-B15-ATCGT-(N)5-(T)20VN-3’;
SEQ ID NO: 4, i.e., 5’-B15-TCATG-(N)5-ME’-3’;
SEQ ID NO: 5, i.e., 5’- B15-ATCGT-(N)5-(G)3-3’; or
SEQ ID NO: 6, i.e., 5’- B15-ATCGT-(N)5-(T)3-3’; or
a nucleotide sequence having 95% identity to any of the above sequences.
Tang teaches designing a set of molecular barcodes to enable the sequencing of many libraries in parallel for Illumina sequencing (Abstract) comprising ATCGT (8 Barcoded and Pooled Libraries GUBZ), and that the GUBZ library comprising said barcode was able to assign 96.62% of reads to a barcode (Pooling of Illumina Libraries with Molecule Barcodes) with relative even number of reads for each unique barcode and quality Illumina reads (Conclusion; see also Read Distribution of Barcoded Libraries), wherein the barcodes in the GUBZ pool comprising ATCGT had a tighter spread of read distributions the two barcode libraries chosen (Read Distribution of Barcoded Libraries).
Tang teaches the importance of pooling and barcoding as the total number of reads and raw bases per lane increases (Pooling of Illumina Libraries with Molecular Barcodes).
EURx teaches an anchored oligo(dT)VN primer that is a string of 20 thymidine residues followed by dV and then dN at the 3’ end, wherein the variable end sequence is anchored to the 5’ end of the poly(A) tail of mRNA, which prevent priming within the poly(A) tail (Description, para 1). EURx teaches that oligo(dT)VN primers may provide an advantage over standard oligo(dT) primers when generating cDNA from poly(A)+ RNA and is recommended for use in cDNA labeling protocols (Description, para 2).
Therefore, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have first substituted the barcode of Steemers for the barcode of Tang as it was taught to be useful for the same purpose and/or motivated by at least the high read assignment of the GUBZ pool barcodes, wherein selecting among that pool would have been obvious to try with a finite number of identified barcodes that were predictable for use in such oligonucleotides as they were demonstrated useful in Tang and wherein the artisan would have desired to address the recognized problem of the increasing number of total numbers of reads and raw bases per lane by choosing appropriate barcodes/barcoding systems in such adapters, as taught by Tang, and also as suggested by Steemers. There would have been a strong expectation of success as Tang is also directed to barcodes for sequencing and such a substitution would have been predictable as a combination of known elements with established functions.
It is further noted that it would be considered, within the broadest reasonable interpretation, that a 1 nt difference in a 47 nucleotide sequence of SEQ ID NO: 1, e.g., utilizing a 6 nucleotide UMI rather than the claimed 5 nucleotide interpreted UMI within the claimed sequences (see instant pg. 14, para 1-2), such that 5’-B15-ATCGT-(N)6-(T)20VN-3’ with a sequence length of 48 would be within 95% sequence identity of 5’-B15-ATCGT-(N)5-(T)20VN-3’.
Also, it would have been obvious to the POSITA before the EFD to have substituted the poly(T) of Steemers for the poly(T)VN of EURx, motivated by the advantage of anchoring to the end of the poly(A) tail, as taught by EURx. There would have been a strong expectation of success as EURx is also directed to sequences capable of priming and such a substitution represents the application of a known technique to improve a known product with established function such that there would be no unexpected outcome.
Additionally, as an alternative to the substitution for A14ME as discussed in the rejection above, it would have been obvious to the POSITA before the EFD to have substituted the A14 of Steemers for the B15 utilized as the alternative adapter sequence, as such was taught as an equivalent sequence for the same purpose by Steemers. See MPEP 2144.06 regarding substituting equivalents known for the same purpose. Thus, a sequence with at least 95% identity to sequence of SEQ ID NO: 1 is obvious as part of the adapter of Steemers.
It further would have been obvious to the POSITA before the EFD of the claimed invention to have appended at least one additional annealing/primer sequence, i.e., a “Seq-1 primer”, as a tag, in the combinatorial indexing/duplication of parts additions for the same reasons as discussed above, wherein in so far as the claims may require a second analyte barcode and a second UMI, such also would be obvious to duplicate for the same reasons.
There would have been a strong expectation of success as each of these additional elements is taught by Steemers and such would have been predictable as routine design choices in combining these known elements according to established functions with no unexpected outcomes.
Response to Arguments
Applicant's arguments filed 06/29/2026 have been fully considered but they are not persuasive.
Regarding the 101 rejection(s), Applicant argues on pg. 6-8, that the claims 13-16 are not directed to non-statutory subject matter and are directed to patent-eligible subject matter. Applicant argues that the claims do not describe a natural product and, as amended, are directed to a composition of matter comprising an amplification primer sequence comprising an RT primer adapted for reverse transcription. Applicant argues that this recites additional elements that integrate the patent-ineligible concept into a practical application. Applicant further argues that any natural product would further be integrated into a practical application. Applicant notes that the claims require that the adapter(s) comprise the same amplification primer sequence that is adapted for RT for the practical application of co-amplification of both DNA and RNA in the same cell, and asserts that the claims are not, therefore, directed to a natural product.
These arguments are not persuasive. As addressed in the updated 101 rejection, the claims remain directed to a set of sequences that encompass, under the broadest reasonable interpretation, a set of sequences that encompass those that may be found in natural genomes, for example. It is noted that an amplification primer sequence adapted for RT may encompass, for example, a random hexamer primer (i.e., any set of six bases) which is frequently used in RT and would be capable to priming both DNA and RNA. As discussed in the rejection, a 6-nucleotide UMI may similarly be any region of 6 nucleotides; where two adapters are required in the set of a de novo-acquired SNP in such a region could represent a UMI. Similarly, the analyte barcodes remain unconstrained and may represent a single nucleotide (e.g., Steemers teaches barcodes may have a length of 1 in para [0056]). The claims recite “comprises” and, as such, do not require a particular order beyond what is specifically claimed, and have a BRI applied for “oligonucleotide adapter” lengths for the separation of such sequence elements that may exist within natural counterparts, such as a genome.
It is further noted that intended use is not sufficient to integrate a natural product into a practical application and that no definition was identified for sequences required to adapt such amplification primer sequences for co-amplification. It is also noted that, while Step 2B was not discussed, MPEP 2106.05(I) recites:
An inventive concept "cannot be furnished by the unpatentable law of nature (or natural phenomenon or abstract idea) itself."…Instead, an "inventive concept" is furnished by an element or combination of elements that is recited in the claim in addition to (beyond) the judicial exception, and is sufficient to ensure that the claim as a whole amounts to significantly more than the judicial exception itself.
As such, the Applicant is encouraged to consider amending to include specific “man-made” sequences or elements that are not found in natural counterparts.
Conclusion
No claims are allowed.
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure:
Thermo Fisher (A new method to remove DNA: Thermo Fisher Scientific [Internet]. Thermo Fisher US; 2020 [cited 2026 Sept 3]. Available from: https://web.archive.org/web/20200805163459/https://www.thermofisher.com/us/en/home/references/ambion-tech-support/rna-isolation/tech-notes/a-new-method-to-remove-dna.html) teaches amplifying both RNA and DNA with an oligo d(T) in an RT-PCR (Fig. 1): “none of these RNA isolation methods produce DNA-free RNA”.
Thus, the artisan would understand an oligo d(T) primer to be adapted for RT of an RNA and adapted for both DNA and RNA co-amplification.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Emma R Hoppe whose telephone number is (703)756-5550. The examiner can normally be reached Mon - Fri 11:00 am - 7:00 pm.
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/EMMA R HOPPE/Examiner, Art Unit 1683
/NANCY J LEITH/Primary Examiner, Art Unit 1636
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