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and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114.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 01/14/2026 has been entered.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Status of Claims
Applicant’s amendment filed 01/14/2026 is acknowledged. Claim 1 has been amended. Claims 2-10 and 19 have been cancelled. Claims 1, 11-18, and 20-25 are pending in the instant application, claims 11-18 and 22-25 remain withdrawn, and claims 1 and 20-21 are the subject of this non-final office action.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 Any rejections or objections not reiterated herein have been withdrawn in light of amendments to the claims or as discussed in this office action.
Previous Rejection
Status of Prior Rejections/Objections:
The objection to claim 1 is withdrawn in view of the amendments to the claim
The prior art rejection under 35 USC 102 directed to claim 1 as being anticipated by Gantt is withdrawn in view of the amendments to the claim.
The prior art rejection(s) under 35 USC 103 as being unpatentable over Gantt in view of Illumina and MGH or Gantt in view of Illumina and MGH and further in view of Slatter are withdrawn upon further search and consideration. See new art rejections over Gantt in view of Illumina and Illumina – Nextera.
Claim Objections
Claim 1 is objected to because of the following informalities:
Claim 1: On pg. 3, in the second paragraph that begins with “and with the proviso that none”, the phrase “or fourth, index sequence” should not have a comma after “fourth”.
Appropriate correction is required.
New Ground(s) of Rejections
Claim Rejections - 35 USC § 103
Claim(s) 1 and 20-21 is/are rejected under 35 U.S.C. 103 as being unpatentable over Gantt (WO 2020/176362 A1; published 09/03/2020; as cited in the IDS dated 07/05/2022) in view of Illumina (Illumina. Indexed Sequencing Overview Guide; 2020 [Internet]. [accessed 2025 Mar 13]. Document #15057455 v08. Available from: http://web.archive.org/web/20210819174703/https://support.illumina.com/content/dam/illumina-support/documents/documentation/system_documentation/miseq/indexed-sequencing-overview-guide-15057455-08.pdf) and Illumina – Nextera (Illumina. Nextera XT DNA Library Prep Kit; 2018 [Internet]. [accessed 2026 Aug. 13]. Document #15031942v03. Available from: https://genome.med.harvard.edu/documents/libraryPrep/IlluminaNexteraXTProtocol.pdf).
Regarding claim 1, Gantt teaches a set of blockers may comprise 9, 10, or more than 10 blockers (para [00129]). Gantt teaches that the sets of blockers may prevent adapter-adapter interactions in sets of massively parallel sequencing library molecules (para [00117-118]; Title; [0099]see para [00192]).
Gantt teaches the region external to the index in the adapters may serve as universal primer binding sites to facilitate simultaneous amplification of all members of a barcode primer library or a subpopulation of members (para [00113]), i.e., “extension primer binding sites”.
Gantt teaches embodiments of sets of blockers that hybridize to regions (from 5’ to 3’ relative to the top strand) upstream of a first index, a pair of adjacent split blockers that hybridize between the first index and an insert, a pair of adjacent split blockers that hybridize between the insert and a second index, and a blocker that hybridizes downstream of the second index, wherein the each of the blockers do not overlap the index (Fig. 35B and 35C; see also Fig. 35A), and wherein the same blocker may be used for the Y-stem annealing/insert-adjacent portion (para [0085]; e.g., Fig. 35B N).
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Gantt teaches that it can be beneficial to design blockers that do not target the index sequence because such “universal” blockers that are index independent allow a minimum number of blockers to be used regardless of the number of different index sequences employed (para [00140]).
Gantt teaches that optimization of such sets of blockers following design comprises replacing top blockers for bottom blockers and vice versa (para [00129]).
Gantt teaches the staggered configuration of blockers from 5’ to 3’ (relative to the top region) complementary to the top region upstream of a first index, the bottom region downstream of the first index and upstream of an insert, the top region downstream of the insert and upstream of a second index, and the bottom region downstream of the second index (Fig. 40F; para [0091]; see also Fig. 40A and Fig. 37E), wherein the figure has been annotated with the relative positions of the claimed blockers to their respective sequencing library molecules for clarity.
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Gantt teaches inserting non-canonical nucleic acids into blockers to increase the blocker’s Tm and optimizing a particular targeted Tm based on other experimental parameters, including salt concentration and additives (para [00129] and [00131]), in particular to ensure the blockers have the desire effect and do not anneal with other adapters (para [00120]). Gantt teaches setting a minimum Tm for blockers (para [0008]).
Gantt teaches that populations of blockers may have minimal or undesired effects, such as daisy-chaining or sequestering other blockers (para [00120]).
Gantt teaches use with different commercially available adapter systems comprising different stems, such as standard dual barcode adapter system T and transposase adapter system N (para [00344]).
Regarding claim 20, Gantt teaches that the blockers may be any length, depending on the size of the adapter or hybridization Tm, including 20-50 bases in length (para [00130]). Gantt teaches an 18 bp sequence (para [00354; Table 17]). Gantt teaches that a blocker may comprise more than 30% nucleobase analogs (para [00129]).
Regarding claim 21, Gantt teaches kits for target enrichment (para [00325]) and sequencing (para [00337]) and using with commercial adapter systems during enrichment/capture (para [00346]).
While Gantt teaches the positioning and complementarity of the hybridization blockers of (i) – (ix) in Fig. 40F; the splitting adjacent blockers in the regions of covered by (ii)-(iii), (iii)-(iv), (vi)-(vii), (viii)-(ix); and sets of blockers that do not overlap index sequences, it fails to explicitly teach the combination of these and that:
each comprise a plurality of Tm increasing nucleotide analogs;
the split blockers are adjacent with 0 nt gap;
(iii) is complementary to a second TE and a complement of a first TE;
that the regions of covered by (ii)-(iii), (iii)-(iv), (vi)-(vii), (viii)-(ix) may be considered sequencing primer-binding sites or complements thereof;
a combination of at least four pairs of split blockers, i.e., (ii)-(iii), (iii)-(iv), (vi)-(vii), (viii)-(ix), with two directed to sequencing primer binding sites and two directed to TEs adjacent to sequencing primer binding sites; and
providing the set of hybridization blockers in a kit.
Illumina teaches a dual-indexed sequencing system (pg. 2, Dual-Index Sequencing Overview, spanning pg. 8) in which the regions between the i5 [i.e., first] index and the insert and the i7 [i.e., second] index and the insert function as sequencing primer binding sites on the same strand (pg. 5), such that the complementary strand would be understood to contain the respective complementary regions (Fig. 3).
Illumina teaches that sequencing primers in TruSeq v3 kits are not compatible with most Nextera libraries (pg. 8, Additional Primers for the TruSeq Cluster Kit v3). Illumina teaches TruSeq commercial kits (pg. 8).
Illumina – Nextera teaches a dual-indexed sequencing system in which the regions between the index and the insert have been added by the transposome [i.e., a transposase] and function as, respectively, Read 1 or Read 2 sequencing primers, and that the transposon end regions are different (pg. 15). Illumina – Nextera teaches Nextera XT and Nextera DNA commercial kits (Table 1).
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 optimized the split blocker embodiment of Fig. 35B such that D and Q bound to the top strand, as such would be obvious to try. Gantt teaches the recognized problem of blockers having minimal or undesired effects such as daisy-chaining or sequestering other blockers and the need to prevent adapter-adapter interactions by using blockers and optimizing blocker binding by trying the finite, identified, and predictable solutions of switching blockers from the top to bottom or bottom to top. Such would have had a reasonable likelihood of success at least because Gantt teaches binding of blockers, that were not split, in such a configuration in Fig. 40F.
Having arrived at the set of blockers targeting sequences of sequencing library template molecules in the same positions as (i) – (ix), it further would have been obvious to the POSITA before the EFD of the claimed invention to have utilize a set of blockers wherein none comprise the first or second insert sequence or complement thereof, motivated by the desire to make a set of universal blockers that is index independent to reduce the number of blockers required, as taught by Gantt.
It also would have been obvious to the POSTIA before the EFD of the claimed invention to have utilized a plurality of Tm increasing nucleotides, wherein the number of Tm increasing nucleotides is identified as a results effective variable, motivated by the desire to optimize the binding temperature of the blocker oligos to achieve the taught minimums and to prevent undesired interactions with other blockers and/or achieve desired binding under chosen experimental conditions, as taught by Gantt.
Given illustration of the split blockers of Gantt as adjacent to one another and use of connected (i.e., unsplit) blockers in alternative embodiments, e.g., Fig. 40F, targeting the internal regions between the indices and the insert, it would have been obvious to the POSITA before the EFD of the claimed invention to design adjacent split blockers separated by 0 nt, wherein the positioning and distance between the blockers is determined to be a routinely optimized variable, as Gantt teaches optimization of the length of the blockers based upon the size of the adapter and the Tm, which also has been discussed above, wherein the relative position of the nucleotide sequence would be considered a range.
At least because Gantt teaches utilizing sets of blockers with different commercially available adapter systems including “dual barcode adapter system T” and “transposase adapter system N”, it would have been obvious to the POSTIA before the EFD of the claimed invention to have designed the set of blockers to be compatible with dual barcode adapter system TruSeq and transposase adapter system Nextera as both are commercially available systems, as obvious species of the genus of commercially available dual barcode adapter systems and transposase adapter systems, respectively, wherein the corresponding template molecule regions between the index and the insert would be, respectively, to which the blockers would be designed to correspond would be two sequencing binding sites on the top strand/complements of the sequencing binding sites on the bottom strand and a two regions that may be interpreted to be a transposon ends flanking an insert that also serve as a sequencing primer binding site, as taught by Illumina and Illumina – Nextera, respectively. And, further motivated by the teachings of Illumina that the sequencing primers of TruSeq and Nextera are generally incompatible, i.e., different sequences, it would have been obvious to the POSITA before the EFD of the claimed invention to have designed each different paired split blockers, including different blockers for the regions directly flanking the insert for the Nextera blockers while, comprising the claimed sequences directed to the pairs of split segments for the reasons above, and to have combined these four pair split blockers in a set of at least 9 blockers, as taught by Gantt, motivated by desire to combine equivalents for the same purpose of avoiding adapter-adapter interactions during sequencing library prep, as taught by Gantt.
Finally, it would have been obvious to the POSITA before the EFD of the claimed invention to have provided the blockers with the sequencing and/or targeting enrichment kits as both are taught for the same purpose of producing nucleic acid sequencing data, as taught by Gantt, and/or to increase the convenience of the blockers for the user.
There would have been a strong expectation of success of each such modification as Gantt teaches the blockers, positioning, means of optimizing, and Illumina and Illumina – Nextera teaches commercially available template molecules sequence regions for use in the design, according to Gantt.
It is further noted that while the order of the regions the plurality of the nucleic acid sequencing library molecules is taught or suggested in the art, the claimed blockers, which are not claimed to span regions do not require the order of the claimed sequencing library molecules. Likewise, the structure of the sequences the may be primer binding sites may be broadly interpreted to encompass a wide range of sequences.
Response to Arguments
Applicant's arguments filed 01/14/2026 have been fully considered but they are not persuasive.
Regarding the arguments directed to the 103 rejections, Applicant argues that the amended claims emphasize a set of nine hybridization blockers including 4 pairs of split hybridization blockers which are directed to two different types of nucleic acid sequencing library molecules. Applicant notes that Example 1 of the instant disclosure describes results obtained with the set of hybridization blockers with two types of nucleic acid sequencing library molecules (TruSeq and Nextera) and said types combined, finding that the Applicant’s nine oligo blocker set in Example 1 blocks approximately equally for the duplexed TruSeq and Nextera compared to the individual single plex libraries (e.g., Fig. 4C).
Applicant argues that none of the cited references teach or suggest all aspects of the present claims, and that no prima facie case of obviousness has been established.
In view of the amendments and upon further search and consideration, the rejection has been updated and clarified. Gantt teaches a set of at least 9 blockers directed to commercial sequencing library adapters, including genera that may include the obvious species TruSeq and Nextera/Nextera XT, wherein it would have been obvious to design toward the taught respective sequence primers and transposon ends. Gantt teaches splitting the blockers in the regions between the index and insert (e.g., Fig. 35B) and producing blockers with sequences of each the regions claimed (e.g., Fig. 40F), wherein it would have been obvious to avoid the index sequences for the reasons discussed above (e.g., to reduce the number of blockers required), and to have include Tm increasing nucleotide analogs and to have optimized position such that the split blocker pairs would have been spaced adjacent by 0 nt, as discussed above, as the number of Tm analogs, the Tm, and length are optimized, as taught by Gantt.
Applicant's arguments fail to comply with 37 CFR 1.111(b) because they amount to a general allegation that the claims define a patentable invention without specifically pointing out how the language of the claims patentably distinguishes them from the references.
Where Applicant emphasizes that in the new claims, the blockers are directed to the claimed sequencing library molecules, it is noted that a recitation of the intended use of the claimed invention must result in a structural difference between the claimed invention and the prior art in order to patentably distinguish the claimed invention from the prior art. If the prior art structure is capable of performing the intended use, then it meets the claim.
In the instant case, the blockers are not claimed to span across regions such that the order would require a particular structure of the blockers nor are the sequencing library molecules tied to particular, for example, SEQ ID NO.
While the data on the blocking is appreciated, Applicant does not recite a comparative ability to block sequencing library molecules in the rejected claims.
Conclusion
No claims are allowed.
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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