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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 11/13/2025 has been entered.
Applicant’s arguments and amendments have been thoroughly reviewed and considered. Claim 2 has been canceled. Claims 1, 3, 6-10, and 12-21 are pending and are examined on the merits herein.
Response to Applicant’s Arguments and Amendments
Regarding the 35 USC 103 Rejections presented in the Final Rejection mailed 8/18/2025, Applicant argues that the newly amended claim 1, which includes the explicit use of a mixture of oligo dT and randomer primers, is an improvement upon Applicant’s existing SMART-Seq technology. Specifically, Applicant states the alleged benefits of the invention noted by Figures 16-18, including enhanced library prep, increased read coverage, and improved library yield (Remarks, page 7). Applicant states that Linnarson teaches their RNA complementary sequences can comprise oligo dT sequences or random sequences, but does not teach a mixture of both as claimed (Remarks, page 8), and thus, claiming that this teaching reads on the instant invention is utilizing impermissible hindsight. Applicant then discusses allegedly unexpected results associated with the use of their invention, as Linnarson and Grunenwald do not suggest the improved results shown by Applicant.
Regarding the claimed results described by Applicant, Figure 16 is a schematic of cDNA synthesis, and does not display any specific data (see para. 54 of the instant specification). While it is possible that the advantages described by Applicant are supported by data, this figure alone is not convincing that the SMART-Seq Plus method provides enhanced library preparation compared to the SMART-Seq method. Regarding Figure 17, Applicant states that this figure shows improved read coverage of 5’ ends and GC-rich regions of a library. It is noted that this data comes from nuclei, and so is specific to at least one cell organelle. The figure itself does not appear to directly address GC regions, though it does state that SMART-Seq Plus has enhanced representation of 5’ regions of gene bodies compared to SMART-Seq. However, it appears that the values near the 5’ end of the gene bodies are incredibly similar between the SMART-Seq group (represented by the triangles) and the SMART-Seq Plus group (represented by the diamonds), and no specific values or statistics are provided to support Applicant’s assertions. In addition, it appears that coverage is generally higher for the SMART-Seq group compared to that of the Smart-Seq Plus group in the middle of sequences. Regarding Figure 18, for nuclei, SMART-Seq Plus does appear to have increased nuclei counts and numbers of genes detected compared to SMART-Seq. However, none of these figures are discussed in any significant detail in the instant specification, and so the specifics of these methods must be based on how “SMART-Seq Plus” is defined in the instant specification.
Para. 79 defines “SMART-Seq Plus” as “a method of preparing cDNA from RNA using a first strand synthesis primer comprising a first amplification primer binding site, a randomer comprising a first amplification primer binding site, and an oligonucleotide switching oligonucleotide that is partially complimentary to the first strand of the cDNA and comprises a second amplification primer binding site. In some embodiments, the first strand synthesis primer comprises an oligo(dT) portion,” (emphasis added). Comparing this definition to the claimed method, it is noted that an oligo(dT) portion of the primer is not required, the first strand synthesis primer does not appear to require all the components listed in instant claim 1 (i.e. the tag and its associated segments), and the claims do not appear to require a template-switching oligonucleotide as stated in the definition. The claim also requires tagmentation that is not clearly included in SMART-Seq Plus methods. Thus, the SMART-Seq Plus method does not appear to exactly align with the claimed method.
Therefore, it is overall unclear if the results described by Applicant are commensurate in scope with the claimed invention, and furthermore, it is unclear if the alleged improvements of SMART-Seq Plus described by Applicant are all present based on the provided figures and remarks.
Additionally, MPEP 716.02(b) I notes that, “The evidence relied upon should establish "that the differences in results are in fact unexpected and unobvious and of both statistical and practical significance." Ex parte Gelles, 22 USPQ2d 1318, 1319 (Bd. Pat. App. & Inter. 1992) .” Applicant states that using a mixture of oligo(dT) and randomer primers provides unexpected results partially because it improves library yield (Remarks, page 11, para. 1). Further search and consideration of such primer mixtures has found that said mixture of oligo(dT) and randomer primers is known in the prior art, as evidenced by Bergtsson et al. (US 2010/0216194 A1) – this reference is cited in the new grounds of rejection below. In this reference, it is stated that “A combination of random hexamer and oligo(dT) priming ensures a high cDNA yield,” (para. 59). Para. 48 of the reference describes putting this primer mixture into practice, and the data is shown in Figure 3B. A combination of oligo(dT) and random hexamer primers were used, and it was concluded that this combination generally leads to an increased yield compared to using oligo dt or random hexamer sequences alone. Thus, the results of an increased yield when using oligo(dT) and randomer primers is not considered unexpected or unobvious (see the full rejection below in the 35 USC 103 Rejections).
Regarding Applicant’s arguments against Linnarson and the claimed primer mixture, the Examiner agrees that para. 51 states that RCS sequences can comprise oligo (dT) sequences or random sequences, but does not appear to use both types of RCS sequences together in a mixture as required by the claim. Therefore, the previous 35 USC 103 Rejections have been withdrawn, for all currently pending claims, but see new grounds of rejection below. It is noted that claim 2 has been canceled, and so this rejection has been rendered moot.
Claim Interpretation
As currently written, in claim 1, the final three lines of the claim read, “wherein the cDNA library comprises a plurality of tagged cDNA fragments comprising the first-read sequencing adapter sequence on a first strand and the second-read sequencing adapter sequence on a second strand,” (emphasis added). As “comprising” is considered open language (see MPEP 2111.03), the inclusion of additional, unclaimed elements in the cDNA fragments is permitted.
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 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.
Claims 1, 3, 6, 10, 12, and 17-20 are rejected under 35 U.S.C. 103 as being unpatentable over Linnarson (US 2012/0010091 A1), in view of Bergtsson et al. (US 2010/0216194 A1), in view of Clarke (WO 2014/029012 A1), in view of Mata (RNA Biology, 2013) and in view of Grunenwald et al. (US 2010/0120098 A1).
Regarding claims 1, 3, 10, 12, and 17-20 Linnarson teaches a method for preparing a cDNA library (Abstract). This method involves:
releasing mRNA from single cells (para. 11)
synthesizing cDNA from the mRNA via a first strand synthesis primer that includes a cell-specific tag (Figure 4 and paras. 11, 16, and 20)
incorporating the tag into the cDNA during synthesis to act as a barcode to identify the cDNA (paras. 11 and 65)
pooling the tagged cDNA samples (para. 11)
amplifying the pooled, tagged cDNA samples to generate a double-stranded cDNA library (para. 11)
sequencing the library (para. 11; instant claim 12)
Linnarson also teaches that the first strand synthesis primer can contain an RNA complementary sequence, and that these sequences can comprise oligo dt primer or random primer regions (para. 51). The first strand synthesis primer can also contain an amplification primer sequence that can be designed as a sequencing primer for downstream analysis, and thus can act as a sequencing adapter sequence (creating a first-read sequencing adapter; para. 67). Adapters are also taught to be added to the cDNA fragments for library sequencing (para. 72). Additionally, the amplification in step e can be done via single primer PCR, which can also be done by using the first strand synthesis primer (para. 67).
However, while Linnarson does teach that the cDNA library originates from a plurality of single cells (paras. 2, 10, and 11) and shows the expression of mitochondrial ribosomal RNA (para. 34), the reference does not specifically teach that the method can be applied to all cell organelles, nor does it mention the elected organelle of cell nuclei. Linnarson also teaches fluorescence activated cell sorting (FACS; e.g. paras. 47 and 128), to separate individual cells, but does not specifically teach the use of the assay to separate the elected organelle of cell nuclei. Linnarson also does not teach that cDNA can be tagged with a unique molecular identifier sequence via a first strand synthesis primer, nor does it teach performing a tagmentation reaction on the pooled, tagged, double-stranded cDNA samples involving a transposome complex. Additionally, as noted above, though Linnarson also teaches that the first strand synthesis primer can contain an RNA complementary sequence, and that these sequences can comprise oligo dt primer or random primer regions (para. 51), these sequences are not necessarily present together in a mixture. Para. 91 also mentions an RNA complementary sequence that is contains an oligo dt with a degenerate anchor oligonucleotide, but this also does not constitute a mixture of oligo dt and random sequences as claimed.
Bergtsson specifically teaches methods for performing RT-PCR on nucleic acids from a cellular sample and performing reverse transcription with first strand cDNA synthesis primers, where said primers are a mixture of oligo dt primers and random primers (Abstract). Para. 48 describes putting this primer mixture into practice, and the data is shown in Figure 3B. A combination of oligo dt and random hexamer primers were used, and it was concluded that this combination generally leads to an increased yield compared to using oligo dt or random hexamer sequences alone. Para. 59 also notes that “A combination of random hexamer and oligo(dT) priming ensures a high cDNA yield.”
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to use the teachings of Bergtsson in Linnarson to have the RNA complementary sequence contain a mixture of oligo dt and random sequences. Linnarson already teaches that either of these sequences could be used for the RNA complementary sequence, providing a reasonable expectation of success that these would both function in the invention of Linnarson. Bergtsson then teaches that the use of a mixture of oligo dt and random sequences can increase cDNA yields, which would motivate the ordinary artisan to use both in the method of Linnarson.
Clarke, in the same field of endeavor, teaches a method of isolating biological targets (Abstract). The separation method specifically involves immobilization via antibodies binding to a target (para. 96, Figure 1), where the antibodies can be linked to a solid support such as a bead (para. 70; Figure 2). This method is taught as working on any cellular organelle, which the ordinary artisan would recognize included nuclei (para. 93).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious to use the method of Linnarson in view of Bergtsson on the spatially separated cell organelles of Clarke. The ordinary artisan would be motivated to combine these references because the isolation methods of Clarke are gentle on cell targets, mitigating potential DNA damage, and are simple, fast, and less costly compared to other methods (para. 16). Furthermore, the inventions of both Clarke and Linnarson have medical applications for diagnostic use (Clarke para. 15, Linnarson paras. 3 and 8-9), providing a context under which the ordinary artisan may wish to combine the references. There would be an expectation of success with this combination because Linnarson states that their method is compatible with the use of beads, and details exemplary embodiments of the invention that specifically involve capturing fragments on beads (e. g. paras. 49 and 102). Therefore, the bead separation methods of Clarke would be usable with the overall method of Linnarson in view of Bergtsson (instant claims 17-20).
However, the combination of Linnarson, Bergtsson, and Clarke does not teach the use of a first strand synthesis primer with a unique molecular identifier, or performing a tagmentation reaction on pooled, tagged, double-stranded cDNA samples involving a transposome complex.
Mata, in the same field of endeavor, teaches a method of reverse transcribing RNA into cDNA involving the use of primers with both barcodes and unique molecular identifiers (Figure 1 and caption). The unique molecular identifier is a random nucleotide sequence (page 1408, column 2). On the primer, the multiplexing barcode is placed in the middle of the UMI, so that the UMI sequence is on both the 5’ and 3’ side of the barcode (Figure 1 and caption).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to incorporate the UMIs taught by Mata into the single PCR primer taught by Linnarson, in view of Bergtsson, and in view of Clarke. This would allow for tags that contain organelle-specific identifiers and UMI sequences. Mata teaches that the use of UMI sequences allows “each original cDNA [to be] individually tagged and…distinguished after PCR amplification,” (page 1408, column 2). One of ordinary skill in the art would recognize that this would lead to greater ease and efficiency when identifying particular samples, or pools of tagged samples, later on in the method of Linnarson, in view of Bergtsson, and in view of Clarke. There would be a reasonable expectation of success with this addition of UMI sequences because the creation of random nucleotide sequences is well known in the art, as evidenced by Mata and Linnarson (e.g., Linnarson para. 51), and once the UMI is incorporated into the primer, the primer will still function in the same manner as it did previously – it will just incorporate the additional UMI sequence into the cDNA tags (instant claim 3).
However, neither Linnarson, nor Bergtsson, nor Clarke, nor Mata teach a tagmentation reaction on pooled, tagged, double-stranded cDNA samples involving a transposome complex.
Grunenwald teaches methods for using a transposase and transposon for generating fragmentation and tagging of DNA targets (Abstract). A Tn5 transposase can be used (para. 52; instant claim 10). Specifically, the method involves incubating target DNA with a transposase and transposon that contains a 5’ tag, where the transposase is used to generate a plurality of DNA fragments, and the transferred strand of the transposon is joined to the 5’ end of the target fragments, creating tagged fragments (para. 18). This is exemplified by Figures 2 and 3, and Figure 3 in particular shows that the tags are identical for each transposon. This results in the 5’ end of each strand of target DNA containing a transposon sequence. Grunenwald that their methods can be used for generating fragment libraries that can later be sequenced (para. 17), and also teaches that the tags can contain sequencing domains (para. 21), therefore acting as sequencing adaptors. The reference also teaches that cDNA can be used in their methods (para. 17).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to add the specific transposome complexes and associated method of Grunenwald to the method of Linnarson, in view of Bergtsson, in view of Clarke, and in view of Mata. Linnarson does teach a method of further processing the cDNA library via fragmentation with DNase (e.g., para. 72), but the ordinary artisan would recognize that using the tagmentation method of Grunenwald would allow for more efficient further processing, as this method is taught by Grunenwald to not require specialized instruments, be easier, faster, and require less hands-on time than other methods, require less DNA sample to operate, and can “generate tagged DNA fragments that are qualitatively and quantitatively representative of the target nucleic acids in the sample from which they are generated,” (para. 16). As Grunenwald can operate on DNA and create a fragmented library, the teachings of this reference are particularly relevant to the method of cDNA library preparation taught by Linnarson, in view of Bergtsson, in view of Clarke, and in view of Mata. Grunenwald also teaches that their invention has medical applications for diagnostic use and chromosome typing (para. 214), providing a context under which the ordinary artisan may wish to combine the references. One of ordinary skill in the art would have a reasonable expectation of success because by combining these references, the result would be a library of cDNA fragments that are tagged with the tags of Grunenwald at each 5’ end. Thus, every first strand would have a first and second-read sequencing adapter and every second strand would have a second-read sequencing adapter. This is not precluded by the instant claims (see “Claim Interpretation” above), and would still allow for sequencing of the fragments, as well as distinguishing first and second strands from one another, which may be important in analyzing downstream results.
Therefore, claims 1, 3, 10, 12, and 17-20 are prima facie obvious over Linnarson, in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald.
Regarding claim 6, Grunenwald teaches that after tagging and fragmentation, the library can undergo an amplification reaction (para. 22). This would allow for additional products to analyze and therefore has the potential to produce more accurate results.
Therefore, claim 6 is prima facie obvious over Linnarson, in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald.
Claims 7-9 are rejected under 35 U.S.C. 103 as being unpatentable over Linnarson (US 2012/0010091 A1), in view of Bergtsson et al. (US 2010/0216194 A1), in view of Clarke (WO 2014/029012 A1), in view of Mata (RNA Biology, 2013), in view of Grunenwald et al. (US 2010/0120098 A1), and further in view of Whitman et al. (US 2010/0330574 A1).
As detailed above, Linnarson, in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald teaches the method of claims 1 and 6. Linnarson and Grunenwald also teach sequencing (Linnarson para. 11 and Grunenwald para. 17), the references do discuss beads (e.g., Linnarson paras. 49 and 102; Clarke para. 70; Mata page 1412, column 2; Grunenwald para. 109), and Grunenwald teaches amplification involving the addition of sequences to the 5’ end of amplification products (see e.g. Figure 10). However, none of these references teach the requirements for amplification on a solid support as detailed in instant claim 7.
Whitman, in the same field of endeavor, teaches an amplification process which adds a tag sequence to the 5’ end of the amplification products. This tag sequence then hybridizes to a complementary anti-tag sequence which is attached to a capture complex (Fig. 1B, paras. 25 and 43). The capture complex can be a solid support, and so subsequent amplification would occur on said support (paras. 25 and 100, claim 15).
Prior to the effective filing date of the claimed invention, it would it would have been prima facie obvious for one of ordinary skill in the art to modify the library preparation method of Linnarson, in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald by using the amplification method taught by Whitman. One of ordinary skill in the art would be motivated to make this change because the method of Whitman can: “…provide assays that require less optimization of primer concentrations; provide quicker results; have lower non-specific background and higher specific signal when using DNA binding dyes; provide more sensitive detection in general; [and] provide a more perfect representation of product/target concentration…,” (para. 7). This method is therefore faster and more efficient compared to other amplification methods. One of ordinary skill in the art would have a reasonable expectation of success using this method because it is specifically compatible with cDNA that has been converted from RNA (para. 69).
Therefore, the method of claims 6-9 is prima facie obvious over Linnarson, in view of Bergtsson, in view of Clarke, in view of Mata, in view of Grunenwald, and further in view of Whitman.
Claim 13 is rejected under 35 U.S.C. 103 as being unpatentable over Linnarson (US 2012/0010091 A1), in view of Bergtsson et al. (US 2010/0216194 A1), in view of Clarke (WO 2014/029012 A1), in view of Mata (RNA Biology, 2013), in view of Grunenwald et al. (US 2010/0120098 A1), and further in view of Hendrickson (WO 2012/012037 A1).
As detailed above, Linnarson, in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald teaches the method of claim 1.
However, neither Linnarson, nor in view of Bergtsson, nor Clarke, nor Mata, nor Grunenwald teach that the first strand synthesis primer has a double-stranded region, comprises a region capable of forming a hairpin, or comprises a region of RNA.
Hendrickson, in the same field of endeavor, teaches an oligonucleotide adapter that is double-stranded (page 1, line 30), has regions capable of forming a hairpin (page 2, lines 5-7), and that the adapter can be made from RNA (page 9, lines 18-20).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to use the adapter of Hendrickson in place of the first strand synthesis primer taught by Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald. MPEP 2143 I (B) states, “The rationale to support a conclusion that the claim would have been obvious is that the substitution of one known element for another yields predictable results to one of ordinary skill in the art.” The adapter structure is well-known in the art, as evidenced by Hendrickson, and one of ordinary skill in the art would find the use of this adapter predictable as it is specifically taught by Hendrickson as working with library preparation methods (page 8, lines 3-5), and that this adapter can be used “under any condition in which sequencing or amplification or both is desirable,” (page 8, lines 24-26).
Therefore, the method of claim 13 is prima facie obvious over Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, in view of Grunenwald, and further in view of Hendrickson.
Claims 14-16 are rejected under 35 U.S.C. 103 as being unpatentable over Linnarson (US 2012/0010091 A1), in view of Bergtsson et al. (US 2010/0216194 A1), in view of Clarke (WO 2014/029012 A1), in view of Mata (RNA Biology, 2013), and in view of Grunenwald et al. (US 2010/0120098 A1), and further in view of Srinivasan et al. (US 2014/0274740 A1).
As detailed above, Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald teaches the method of claim 1.
However, neither Linnarson, nor in view of Bergtsson, nor Clarke, nor Mata, nor Grunenwald teach that the first strand synthesis primer is attached to a bead, and that tagged cDNA samples are synthesized on said bead. These references also do not teach that each cell organelle is encapsulated by a droplet with the bead before initial mRNA release, or that the droplet is generated using a droplet actuator. Though Clarke does teach separating cell organelles, as described above, this does not occur in droplets.
Srinivasan, in the same field of endeavor, teaches a method of library preparation that can be used with cDNA copied from mRNAs (para. 216) where oligonucleotide primers are attached to fragmented target DNA, and then fragments are attached to capture beads before amplification (para. 245). This PCR amplification occurs within droplets on the beads, and beads are captured in wells (para. 245). This method is taught within the context of 454 sequencing. Para. 212 of this reference also teaches that droplets can be generated using a droplet actuator.
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to substitute the synthesizing, pooling, and amplifying steps of the method taught by Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald with that of Srinivasan. MPEP 2143 I (B) states, “The rationale to support a conclusion that the claim would have been obvious is that the substitution of one known element for another yields predictable results to one of ordinary skill in the art.” The method of using droplets and beads is well-known in the art, as evidenced by Srinivasan, and the ordinary artisan would find the results of this method predictable as it is taught in Srinivasan to work for any nucleic acids in a test sample, and the reference clearly teaches the use of cDNA copied from mRNA (paras. 216 and 245).
Therefore, the method of claims 14-16 are prima facie obvious over Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, in view of Grunenwald, and further in view of Srinivasan.
Claims 21 are rejected under 35 U.S.C. 103 as being unpatentable over Linnarson (US 2012/0010091 A1), in view of Bergtsson et al. (US 2010/0216194 A1), in view of Clarke (WO 2014/029012 A1), in view of Mata (RNA Biology, 2013), in view of Grunenwald et al. (US 2010/0120098 A1), and further in view of Sigurgeirsson et al. (PLoS ONE, 2014).
As detailed above, Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald teaches the method of claim 1.
However, none of these references detail 3’ tag counting.
Sigurgeirsson teaches methods of 3’ tag counting to improve expression measurements in a sample, particularly when quality of said sample is low (Abstract). Sigurgeirsson specifically discusses the use of RNA, and notes that when RNA is fragmented, such as what occurs during library preparation, a range of quality and degraded RNA sequences are created (page 1, column 2, para. 3). These differences in general quality can lead to false positive expression results during analysis (page 2, column 2). Sigurgeirsson performed library preparation and sequencing, and then 3’ tag counting was done, which involved downstream computational analysis (Figure 1). Sigurgeirsson concluded that using 3’ tag counting is simple and straightforward, prevents false positives, and allows degraded sample to be useable for analysis (page 6, column 2, para. 3). The false positive benefits in particular are relevant for even higher quality samples. The caption of Supplementary Figure 1 makes it clear that though RNA was used, it is cDNA that is eventually present in the libraries examined (page 9).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to use the 3’ tag counting analysis methods of Sigurgeirsson in the method of Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald. Once a library is prepared and sequencing is performed, 3’ tag counting can be done entirely analytically, and therefore requires no additional lab work or reagent resources. Sigurgeirsson details the many benefits of using this method that would be motivating to the ordinary artisan, including being able to use lower quality samples and elimination of false positives in analyses. As Sigurgeirsson clearly lays out the results for using this method on RNA (and resulting cDNA libraries) post sequencing, and Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, and in view of Grunenwald teach library sequencing, there would be a reasonable expectation of success.
Therefore, claim 21 is prima facie obvious over Linnarson, in view of in view of Bergtsson, in view of Clarke, in view of Mata, in view of Grunenwald, and in view of Sigurgeirsson.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The filing of a terminal disclaimer by itself is not a complete reply to a nonstatutory double patenting (NSDP) rejection. A complete reply requires that the terminal disclaimer be accompanied by a reply requesting reconsideration of the prior Office action. Even where the NSDP rejection is provisional the reply must be complete. See MPEP § 804, subsection I.B.1. For a reply to a non-final Office action, see 37 CFR 1.111(a). For a reply to final Office action, see 37 CFR 1.113(c). A request for reconsideration while not provided for in 37 CFR 1.113(c) may be filed after final for consideration. See MPEP §§ 706.07(e) and 714.13.
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Claims 1, 6-9, 12-15, and 21 are rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-18 of U.S. Patent No. 10,975,371 B2 in view of Clarke (WO 2014/029012 A1).
Claim 1 of the ‘371 patent recites all of the limitations of instant claim 1, with the exception of using single cells rather than single cell organelles. It is noted that in the ‘371 patent, the first synthesis primer is said to contain a “different second portion,” which can be analogous to the UMI of the instant first synthesis primer. This is because each tag has a different second portion, and therefore each portion would be unique. Regarding the use of single cells versus cell organelles, Clarke teaches a method of isolating biological targets (Abstract). The separation method specifically involves immobilization via antibodies binding to a target (para. 96, Figure 1), where the antibodies can be linked to a solid support such as a bead (para. 70; Figure 2). This method is taught as working on any cellular organelle (para. 93). Thus, it would have been obvious to use the method of the ’371 patent on the spatially separated cell organelles of Clarke. The isolation methods of Clarke are shown to be gentle on cell targets, mitigate potential DNA damage, and are simple, fast, and less costly compared to other methods (para. 16). Furthermore, Clarke has medical applications for diagnostic use (Clarke para. 15), providing a context under which the ordinary artisan may wish to combine the references.
Claim 2 of the ‘371 patent recites the same limitation as instant claim 6, and so reads on this claim.
Claim 3 of the ‘371 patent depends on claim 2 and recites adding an additional sequence to the amplified, tagged cDNA fragments, thus also reading on instant claim 6.
Claim 4 of the ‘371 patent depends on claim 3 and recites that this additional sequence is a primer binding sequence for hybridization on a solid support and performing amplification on said solid support. Claim 4 thus meets all of the limitations of instant claim 7.
Claims 5-6 of the ‘371 patent depend on claim 4 and recite the same limitations as instant claims 8-9, and thus read on these claims.
Claim 7 of the ‘371 patent recites sequencing the tagged cDNA fragments, and thus reads on instant claim 12.
Claim 8 of the ‘371 patent recites performing sequencing via 3’ tag counting, and so reads on instant claims 12 and 21.
Claim 9 of the ‘371 patent recites performing sequencing via whole transcriptome analysis, and so reads on instant claim 12.
Claims 10 and 11 of the ‘371 patent recite the limitations of option a of instant claim 13, and so read on this claim.
Claims 12-14 of the ’371 patent depend on claim 10 and recite options b-d of instant claim 13, thus also reading on this claim.
Claim 15 of the ‘371 patent specifically notes a UMI sequence, and thus also reads on instant claim 1.
Claims 16-18 of the ‘371 patent recite the limitations of instant claims 14-15, and so read on these claims.
Conclusion
No claims are currently allowable.
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/F.F.G./Examiner, Art Unit 1681
/ANGELA M. BERTAGNA/Primary Examiner, Art Unit 1681