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 .
Claim Status
Claims 3-4, 7-8, 19-20, 23-25, 29-30 and 33 have been amended (1/30/2026). Claims 61-66 are new (1/30/2026). No new matter was added. Thus, claims 3, 4, 7, 8, 11, 19, 20, 23-25, 28-30, 33 and 61-66 are under examination (1/30/2026).
Office Action: Notice
Any objection or rejection of record in the previous Office Action, mailed 10/1/2025,
which is not addressed in this action has been withdrawn in light of Applicants' amendments
and/or arguments. This action is FINAL
Election/Restrictions
Applicant’s election without traverse of Group I in the reply filed on August 11, 2025 is acknowledged. Claims 36-38, 41-42 and 44 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to nonelected Groups 2 and 3, there being no allowable generic or linking claim. Further, since Group I was elected, species election I of nucleic acids and species II of three-dimensional region were made. Election was made without traverse in the reply filed on 8/11/2025.
Priority
Claims 3, 4, 7, 8, 11, 19, 20, 23-25, 28-30, 33 and 61-66 receive a priority date of 12/12/2019, the effective filing date of GB Provisional Patent GB1918340.9. All priority documents have been received.
Objections/Deficiencies Withdrawn
Specification:
The objections to the specification due to the use of a trademark or tradenames are withdrawn in view of Applicant’s amendments.
The deficiency for nucleotide and/or amino acid sequences appearing in the listing not being identified by sequence identifiers in accordance with 37 CFR 1.821(a), where sequence identifiers for nucleotide and/or amino acid sequences must appear as a “Sequence Listing”, is withdrawn in view of Applicant’s amendments.
Claims:
The minor grammatical objections to claim 3 are withdrawn in view of Applicant’s amendments.
New Objections
Claim 62 is objected to because of the following informality:
Claim 62 at line 1; “he transcript” should be replaced with “the transcript.”
Rejections Withdrawn
Claim Rejections - 35 USC § 112(b)
The rejection of claims 3-4, 7-8, 11, 19-20, 23-25, 28-30, and 33 under 35 U.S.C. 112(b) or pre-AIA 35 U.S.C. 112, 2nd paragraph, is withdrawn in view of Applicant’s amendments of claims 3, 7, 8, 19, 20, 23-25, 29-30 and 33.
Rejections Maintained
Claim Rejections: 35 USC § 102
Claims 3, 4, 7, 8, 11, 19, 20, 23-25, 28-30 and 33 are rejected under 35 U.S.C. 102 (a)(1) and (a)(2) as being anticipated by Vijayan et al., (US PGPub 2019/0093103 A1, published 3/28/2019).
Regarding claims 3-4, 7-8 and 11, Vijayan teaches a bead or detection probe with a covalently attached chemical compound and a covalently attached DNA barcode and methods for using such beads, where the bead has many substantially identical copies of the chemical compound and many substantially identical copies of the DNA barcode (Abstract). Further, Vijayan teaches that the addition of nucleic acid barcodes to the beads allows the identity of the compound present within the beads to be carried along to the assay volume where very high throughput assays may be performed without needing robotics or spatial indexing of compounds within microtiter plates (Paragraph 7, lines 5-10). Specifically, Vijayan teaches that while incubating cells or tissues under different perturbations within well plates, followed by single-cell analysis and comparisons between transcript profiles can be done, the number of drugs that can be examined is limited by the plate capacity (Paragraph 11, lines 5-10). Vijayan also teaches that the previously described detection probe or bead methodology provides a system for screening chemical compounds, comprising: (a) A picowell array plate comprising a plurality of picowells, wherein each picowell has a top aperture that defines an opening at the top of the picowell, a bottom that is defined by a floor, wherein the top aperture is separated from the floor, and wherein a wall resides in between the top aperture and the floor; (b) A bead or detection probe disposed in a picowell, wherein the bead comprises a plurality of substantially identical bead-bound DNA barcodes, and a plurality of substantially identical bead-bound compounds, (c) Wherein the bead comprises a bead-bound DNA barcode or binding region that takes the form of either a concatenated DNA barcode or an orthogonal DNA barcode, and wherein if the DNA barcode takes the form of a concatenated DNA barcode the concatenated DNA barcode is made by a method that: (i) Uses click chemistry, or (ii) Uses a repeating cycle of steps, wherein the repeating cycle of steps comprises using a splint oligonucleotide (splint oligo) that is capable of hybridizing to a partially made bead-bound DNA barcode, and wherein the hybridizing is mediated by an annealing site on the splint oligo and a corresponding, complementary annealing site in the partially made bead-bound DNA barcode, wherein the annealed splint oligo is used as a template for extending the partially made DNA barcode using DNA polymerase, and wherein the splint oligo contains bases that are complementary to a DNA barcode module that is to be polymerized to the partially made DNA barcode, (d) Wherein each one of the plurality of substantially identical bead-bound compounds comprises one or more chemical library monomers, and wherein each bead-bound DNA barcode module identifies a corresponding chemical library monomer, wherein the term “compound” is used to refer to a completed product that comprises one or more chemical library members, and wherein the completed DNA barcode identifies the compound (Paragraph 12, lines 1-20).
Specifically, Vijayan teaches that the concatenated DNA barcode is coupled to the bead, but is: (i) not coupled to the bead by way of any photocleavable linker or index sequence, (ii) not coupled to the bead by any enzymatically cleavable linker; or (iii ) not coupled to the bead by any kind of cleavable linker (Paragraph 47, lines 1-3). Vijayan also teaches that herein the release-monitor bead comprises a bead, a quencher, a fluorophore, and a photocleavable linker that couples the fluorophore to the bead for illumination, the method comprising, in this order, (i) Providing a resin, (ii) Coupling a lysine linker to the resin, wherein the reagent containing the lysine linker is L-Fmoc-Lys(4-methyltrityl)-OH, (iii) Removing the Fmoc protecting group, (iv) Coupling the quencher using a reagent that is quencher-N-hydroxysuccinimide (quencher-NETS) as the source of quencher, (v) Removing the 4-methyltrityl protecting group using a reagent comprising trifluoroacetic acid, (vi) Coupling a photocleavable linker to the epsilon amino group of lysine, wherein the photocleavable linker is provided by a reagent that is, Fmoc-photocleavable linker-OH, (vii) Coupling the fluorophore (Paragraph 52, lines 1-15).
Vijayan further teaches that the previously described method of spatial barcoding via beads or detection probes can be applied to prepare barcoded mRNA from each sample in isolation and then perform comprehensive RNA profiles for every sample (Paragraph 11, lines 5-10).
Regarding claim 19, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes includes orthogonal barcoding where each individual module gets covalently bound to its own unique attachment site on the bead and where once a module gets attached to a given site on the bead, no further modules will be connected to the module that is already attached (Paragraph 176, lines 15-20).
Regarding claims 20 and 23, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes includes a stepped picowell, a three dimensional structure or physical structure with three compartments and two steps where the top compartment is widest and is configured for accepting cap where most of the top compartment is occupied by the cap in the situation where the picowell is capped and the middle compartment is configured for being occupied mainly by, or solely by, reagents (Figure 29). Further, Vijayan teaches that the reagents can include buffer, enzyme substrates, one or more salts, and a preservative or stabilizer such as dithiothreitol, RNAse inhibitor, glycerol, or DMSO and the lowest compartment is configured for being occupied by bead, that is, a bead with coupled both a DNA library and with releasable compounds (Figure 29; Paragraph 381, lines 1-15).
Regarding claim 24, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes includes cleaving the DNA barcode from the bead before sequencing and can encompass a method where bead-bound DNA barcode is cleaved from the bead, thereby releasing the DNA barcode in a soluble form, prior to amplification, or prior to sequencing, or prior to any type of sequence identification technique such as hybridizing with a nucleic acid probe (Paragraph 401, lines 1-5).
Regarding claim 25, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes includes bead-bound compositions, systems, and methods, that uses at least one splint oligo or split detection probes, at least two splint oligos, at least three splint oligos, at least four splint oligos, at least five splint oligos, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at last 13, at least 14, at least 20 splint oligos, or less than 20, less than 15, less than 10, less than 8, less than 6, less than 4, less than 3, less than 2 splint oligos (Paragraph 291, lines 1-5).
Regarding claim 28, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes can be applied to amplified DNA while attached to a bead and DNA in amplified form is easier to sequence that non-amplified DNA and once single stranded, a splint oligo is added to bridge the ends of the tag DNA, and this is followed by extension and ligation of the splint oligo (Paragraph 407, lines 1-5).
Further, Vijayan teaches that the distance from primer annealing site to a DNA barcode module comprises a polynucleotide including a first nucleic acid that is an annealing site for a sequencing primer, and a second nucleic acid that is a DNA barcode module, the first nucleic acid can be immediately upstream of the second nucleic acid. where, the first nucleic acid can be upstream of the second nucleic acid, where the first and second nucleic acids are separated from each other by one, two, three, four, five, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or more nucleotides, or by about one, about two, about three, about four, about five, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, or about 15 nucleotides (Paragraph 221, lines 1-10).
Regarding claims 29-30, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes includes the option of creating a DNA barcode that includes a terminal nucleic acid that encodes DNA hairpin which includes, at the 3-prime end, a nucleic acid that possesses an annealing site for a sequencing primer, a bend taking the form of about four bases that are not base-paired, and a sequencing primer that is capable of bending around and forming base pairs with the sequencing primer annealing site (Paragraph 242, lines 1-5). Specifically, Vijayan teaches that these optional encoding regions can include: (1) Linkers to attach chemical library member to a substrate, such as a bead; (2) Linkers to attach nucleic acid barcode to a substrate, such as a bead; (3) Cleavable linkers, for example, cleavable by UV light, cleavable by an enzyme such as a protease; (4) Non-cleavable linkers; (5) Bifunctional linkers; (6) Multi-functional linkers; and (7) Plurality of beads used for linking (Paragraph 314, lines 1-5).
Regarding claim 33, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes includes beads or probes where DNA barcodes are mainly attached on the exterior surface; however one reason to NOT make and use beads with internal DNA barcodes, is the low permeation of DNA oligomers to the interior spaces, and low permeation of DNA ligases to interior spaces (ligases for connecting DNA modules to each other to create the finished DNA barcode) (Paragraph 355, lines 1-5). Specifically, Vijayan teaches that the previously described method of spatial barcoding via beads or detection probes can be applied to amplified DNA while attached to a bead and DNA in amplified form is easier to sequence that non-amplified DNA and once single stranded, a splint oligo is added to bridge the ends of the tag DNA, and this is followed by extension and ligation of the splint oligo (Paragraph 407, lines 1-5).
Vijayan teaches each and every limitation of claims 3, 4, 7, 8, 11, 19, 20, 23-25, 28-30 and 33, and therefore Vijayan anticipates claims 3, 4, 7, 8, 11, 19, 20, 23-25, 28-30 and 33.
Applicant’s Response: The Applicant argues that Vijayan does not teach “spatial barcoding”, specifically, illumination-based cleavage used in indexing or iterative cycles of illumination and index addition as recited in steps (c)-(e). The Applicant further argues that Vijayan excludes the use of photocleavable linkers for barcode attachment and only teaches such linkers in connection with fluorophore release.
Examiner’s Response to Traversal: Applicant’s arguments have been carefully and fully considered but are not found persuasive, as discussed below.
As of note, during examination, claims are given their broadest reasonable interpretation (BRI) consistent with the specification (see MPEP 2111). Further, a claim is anticipated when a single reference discloses each and every element, either expressly or inherently, arranged as in the claim (see MPEP 2131, 2112).
Therefore, under BRI, independent claim 3 does not require any specific optical confinement mechanism, photomask, or imaging system, nor does it exclude bead-based or probe-based systems. The recited “spatially barcoding” broadly encompasses barcoding associated with localized positions or sites. Therefore, the Applicant’s assertion that Vijayan does not disclose spatial barcoding is not persuasive. Vijayan teaches spatially organized barcoding systems using beads and detection probes associated with localized structures. For example, Vijayan teaches spatial barcoding via beads or detection probes (Abstract; Paragraph 7, lines 5-10), and further teaches stepped picowells and three-dimensional compartmentalized structures in which beads occupy defined locations (Paragraphs 220-221, 291). These disclosures showcase that Vijayan associates barcodes with discrete localized physical positions, which reasonably meet the claimed spatially barcoding under BRI. Notably, the Applicant’s argument improperly limits “spatially barcoding” to light-patterned tissue regions, which is not required by the claim language.
The Applicant further argues that Vijayan does not teach illumination that cleaves or alters a photocleavable group. This argument is not persuasive. Vijayan teaches cleavable linkers, including UV-cleavable linkers (Paragraph 314). UV-cleavable linkers, are, by definition, cleaved by illumination. Specifically, UV light form is a form of illumination, and therefore such linkers constitute photocleavable groups responsive to illumination. Vijayan further teaches that these linkers are used in connection with nucleic acid barcodes and bead-based systems (Paragraph 314), thereby incorporating illumination-responsive cleavage chemistry into the barcoding framework. Thus, Vijayan expressly discloses the claimed limitation of “illuminating…wherein the illumination cleaves or alters the photocleavable group.” The Applicant’s argument that such linkers are used for a different purpose (i.e., fluorophore release) is not persuasive because anticipation does not require identical purpose, only that the limitation is disclosed (see MPEP 2131).
Additionally, the Applicant argues that Vijayan does not teach repeated cycles of illumination and index addition. This argument is not persuasive. Vijayan teaches iterative barcode construction, including; split oligo-mediated extension and ligation to build DNA barcodes (Paragraph 407), barcode modules and orthogonal barcoding schemes (Paragraph 355), and structured spacing and assembly of barcode regions (Paragraphs 221, 242). These teachings demonstrate that Vijayan performs multi-step, iterative barcode assembly, which under BRI corresponds to repeated steps of activation and index addition.
To the extent that the Applicant argues that Vijayan does not explicitly describe each iteration as preceded by illumination, the rejection relies on inherency. Under MPEP 2112, a limitation is inherent when it is a necessary result of the disclosed system. Vijayan expressly teaches light-cleavable (UV cleavable) linker chemistry (Paragraph 314) and iterative barcode assembly processes (Paragraph 407). When these teachings are read together, repeated cycles of cleavage/activation and barcode addition are a natural and necessary consequence of the disclosed barcoding workflow under the BRI of the claim. Thus, the repetition of steps (c) and (d) are inherently disclosed.
Further, the Applicant relies on a passage stating that certain barcodes are not coupled via cleavable linkers, citing Paragraphs 47 and 52. This argument is not persuasive. Specifically, Paragraph 47 describes a specific embodiment in which a concatenated DNA barcode is not coupled to a bead via a cleavable linker. However, this disclosure is limited to that embodiment and does not constitute a general disclaimed of cleavable linker chemistry. Vijayan elsewhere teaches cleavable linkers, including UV-cleavable linkers (Paragraph 314). Thus, Vijayan as a whole clearly contemplates the use of illumination-responsive cleavable linkers, even if certain embodiments do not employ them for barcode attachment. It is well established that a prior art reference must be considered for all that it teaches, and that a single embodiment lacking a feature does not negate other embodiments that include that feature (see MPEP 2131). Accordingly, the Applicant’s reliance on Paragraphs 47 and 52 is misplaced and does not overcome the rejection.
Accordingly, the rejection of claims 3, 4, 7, 8, 11, 19, 20, 23-25, 28-30 and 33 under 35 USC 102 is maintained.
Notwithstanding the above rejection, the prior art of record, does not clearly disclose illumination that is spatially restricted to a defined region within tissue such that index sequences are added only to detection probes within the illuminated region and correspond to that tissue location. Accordingly, the Examiner recommends amending the independent claim to recite spatially restricted, illumination-controlled indexing.
New Rejections
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 65 is 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.
Claim 65 recites the limitation "the 3’ end" in line 2. There is insufficient antecedent basis for this limitation in the claim.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
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.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claim(s) 61-66 are rejected under 35 U.S.C. 103 as being unpatentable over Vijayan et al., (US PGPub 2019/0093103 A1, published 3/28/2019), as applied to claims 3, 4, 7, 8, 11, 19, 20, 23-25, 28-30 and 33 in view of Nestorova et al. (“Lab-on-a-chip mRNA purification and reverse transcription via a solid-phase gene extraction technique”, Royal Society of Chemistry, published 2017).
As previously described, Vijayan teaches a bead or detection probe with a covalently attached chemical compound and a covalently attached DNA barcode and methods for using such beads, where the bead has many substantially identical copies of the chemical compound and many substantially identical copies of the DNA barcode (Abstract). Further, Vijayan teaches that the addition of nucleic acid barcodes to the beads allows the identity of the compound present within the beads to be carried along to the assay volume where very high throughput assays may be performed without needing robotics or spatial indexing of compounds within microtiter plates (Paragraph 7, lines 5-10). Specifically, Vijayan teaches that while incubating cells or tissues under different perturbations within well plates, followed by single-cell analysis and comparisons between transcript profiles can be done, the number of drugs that can be examined is limited by the plate capacity (Paragraph 11, lines 5-10).
Regarding claims 61-63, Vijayan teaches that the previously described detection probe or bead methodology provides a system for screening chemical compounds, comprising: (a) A picowell array plate comprising a plurality of picowells, wherein each picowell has a top aperture that defines an opening at the top of the picowell, a bottom that is defined by a floor, wherein the top aperture is separated from the floor, and wherein a wall resides in between the top aperture and the floor; (b) A bead or detection probe disposed in a picowell, wherein the bead comprises a plurality of substantially identical bead-bound DNA barcodes, and a plurality of substantially identical bead-bound compounds, (c) Wherein the bead comprises a bead-bound DNA barcode or binding region that takes the form of either a concatenated DNA barcode or an orthogonal DNA barcode, and wherein if the DNA barcode takes the form of a concatenated DNA barcode the concatenated DNA barcode is made by a method that: (i) Uses click chemistry, or (ii) Uses a repeating cycle of steps, wherein the repeating cycle of steps comprises using a splint oligonucleotide (splint oligo) that is capable of hybridizing to a partially made bead-bound DNA barcode, and wherein the hybridizing is mediated by an annealing site on the splint oligo and a corresponding, complementary annealing site in the partially made bead-bound DNA barcode, wherein the annealed splint oligo is used as a template for extending the partially made DNA barcode using DNA polymerase, and wherein the splint oligo contains bases that are complementary to a DNA barcode module that is to be polymerized to the partially made DNA barcode, (d) Wherein each one of the plurality of substantially identical bead-bound compounds comprises one or more chemical library monomers, and wherein each bead-bound DNA barcode module identifies a corresponding chemical library monomer, wherein the term “compound” is used to refer to a completed product that comprises one or more chemical library members, and wherein the completed DNA barcode identifies the compound (Paragraph 12, lines 1-20).
Specifically, Vijayan teaches that the concatenated DNA barcode is coupled to the bead, but is: (i) not coupled to the bead by way of any photocleavable linker or index sequence, (ii) not coupled to the bead by any enzymatically cleavable linker; or (iii ) not coupled to the bead by any kind of cleavable linker (Paragraph 47, lines 1-3). Vijayan also teaches that herein the release-monitor bead comprises a bead, a quencher, a fluorophore, and a photocleavable linker that couples the fluorophore to the bead for illumination, the method comprising, in this order, (i) Providing a resin, (ii) Coupling a lysine linker to the resin, wherein the reagent containing the lysine linker is L-Fmoc-Lys(4-methyltrityl)-OH, (iii) Removing the Fmoc protecting group, (iv) Coupling the quencher using a reagent that is quencher-N-hydroxysuccinimide (quencher-NETS) as the source of quencher, (v) Removing the 4-methyltrityl protecting group using a reagent comprising trifluoroacetic acid, (vi) Coupling a photocleavable linker to the epsilon amino group of lysine, wherein the photocleavable linker is provided by a reagent that is, Fmoc-photocleavable linker-OH, (vii) Coupling the fluorophore (Paragraph 52, lines 1-15).
Vijayan further teaches that the previously described method of spatial barcoding via beads or detection probes can be applied to prepare barcoded mRNA from each sample in isolation and then perform comprehensive RNA profiles for every sample (Paragraph 11, lines 5-10).
Regarding claims 64-66, Vijayan teaches that reducing damage via coupling DNA to beads via their 3 ' – end, where certain chemical transformation may damage exposed 3 ' - hydroxyl groups of nucleic acids and for instance Mitsunobu reactions allow the conversion of primary and secondary alcohols to esters, phenyl ethers, thioethers and various other compounds, which might render exposed 3 ' - ends unreactive to subsequent processing steps, or cause the now modified 3 ' - end to participate in further chemical reactions (Paragraph 310, lines 1-5). Further, Vijayan teaches that in some embodiments , the DNA tags may be attached to beads via their 3 ' - end , so only the 5 ' - end is exposed to solution (Paragraph 310, lines 5-10).
Additionally, Vijayan teaches that the reagents , systems , and methods of the present disclosure encompass bead - bound nucleic acids , such as a bead - bound DNA or a bead - bound DNA tags , where coupling to the bead involves the 3 ' - terminus (or the 3 ' - end) of
the DNA, where ssDNA that comprises a DNA barcode is coupled by way of the 3 ' - end , of the ssDNA, sequencing can be initiated by hybridizing only one sequencing primer, where this sequencing primer hybridizes upstream of the entire DNA barcode , and where this hybridizing is at or near the bead - bound end of the coupled ssDNA (Paragraph 311, lines 1-10). Vijayan also teaches that as an alternative to using only one sequencing primer , a plurality of sequencing primers can be used, where each sequencing primer hybridizes upstream to a particular DNA barcode module (Paragraph 311, lines 1-10). Vijayan further teaches that for example, if a given DNA barcode contains five DNA barcode modules, and where the DNA is coupled to a bead by way of its 3 ' - end, the DNA barcode can include five different primer annealing sites, where each primer anneals given DNA barcode modulating site is located just upstream of a given DNA barcode molecule (Paragraph 311, lines 10-15).
However, Vijayan does not teach or suggest a specific binding region of the detection probe comprised of RNA, specifically mRNA, that is elongated via reverse transcription.
Nestorova teaches extraction and purification of high-quality RNA is a crucial initial step required for a variety of genomic assays, using a solid phase gene extraction (SPGE) method for automated extraction, purification and reverse transcription of mRNA in a microfluidic device (Abstract). Nestorova further teaches that the specificity and RNA loading capacity of the probes were evaluated using conventional qPCR, where this procedure was successfully used to extract, purify, and transcribe mRNA from rat glioblastoma cell spheroids in less than seven minutes and analysis of the product confirmed that the SPGE technique selectively captures and inherently purifies high-quality mRNA directly from biological material with no need for additional pre-processing steps (Abstract). Further, Nestorova teaches that nucleic acid-based assays require high quality RNA from cells and tissues that is free of proteins or inhibitors that interfere with downstream reverse transcription (RT) and PCR amplification and traditional methods for RNA extraction involve multiple steps and rely on tissue homogenization to release the genetic material and eliminate inhibitors (Introduction: Paragraph 1). Nestorova also teaches that this method simplifies the isolation of RNA from tissue or cells to a single pin prick and specifically, solid phase gene extraction (SPGE) was performed with miniature needles functionalized with oligo(-dT) or gene-specific sequences via simply inserting the needle into the specimen, the target RNA is collected on its tip (Introduction: Paragraphs 2-3).
More so, Nestorova teaches that the reported method describes the integrated collection, purification and reverse transcription of mRNA, while eliminating the need for cell lysis and multi-step mRNA purification and the resulting heterogeneous sample of whole mRNA can be analyzed by shotgun sequencing (RNA-Seq) or RT-PCR quantification of one or more gene-specific mRNA sequences where mRNA extraction is achieved using needles with amino-modified dT(15) oligonucleotides immobilized to their surface (Materials and Methods: Paragraph 1; Fig. 1). Further, Nestorova teaches that the specificity and loading capacity of the SPGE needles were evaluated using total rat RNA (20 ng μL−1) as a source of biological material, where the specificity was assessed using electrophoretic sizing of the RNA; the loading capacity was quantified using a RT-qPCR LightScanner®32 via functionalized steel probes (SPGE of mRNA from total RNA sample: specificity and concentration studies: Paragraph 1).
It would have been obvious to one of ordinary skill in the art at the time of the invention to modify the system of Vijayan to incorporate mRNA-binding probes and reverse transcription as taught by the SPGE reference in order to enable gene expression analysis and transcript profiling of biological samples. Both references are directed to nucleic acid analysis of biological samples and share the common objective of identifying and characterizing molecular content within cells or tissues. Vijayan expressly contemplates RNA analysis (Paragraph 11), and Nestorova provides a known and established technique for selectively capturing and reverse transcribing mRNA for such analysis. Therefore, a person of ordinary skill in the art would have been motivated to combine these teachings to enhance Vijayan’s system with known mRNA capture and reverse transcription techniques.
Furthermore, a person of ordinary skill in the art would have had a reasonable expectation of success in making this combination because both references operate within the same technical field of nucleic acid hybridization, amplification, and sequencing, and because reverse transcription of mRNA using oligo(dT) or sequence-specific probes was a well-understood and routine technique at the time of the invention. Vijayan already teaches nucleic acid constructs capable of hybridization and enzymatic manipulation (Paragraph 311), which are directly compatible with reverse transcription workflows, and the SPGE reference demonstrates that mRNA can be captured on probe surfaces and successfully reverse transcribed in situ. Thus, the combination would have predictably resulted in a system in which mRNA bound to a detection probe is elongated via reverse transcription for downstream analysis. Accordingly, it would have been obvious to modify Vijayan in view of Nestorova to arrive at the claimed invention, including a detection probe comprising an RNA binding region that is elongated via reverse transcription.
Conclusions
No claim is allowed.
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 extension fee 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 date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ELIZABETH ROSE LAFAVE whose telephone number is (703)756-4747. The examiner can normally be reached Compressed Bi-Week: M-F 7:30-4:30.
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/ELIZABETH ROSE LAFAVE/ Examiner, Art Unit 1684
/HEATHER CALAMITA/ Supervisory Patent Examiner, Art Unit 1684