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 .
Election/Restrictions
Applicant’s election without traverse of Group III, claims 18-23, in the reply filed on04/09/2026 is acknowledged.
Claims 1 and 13 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim.
Amendments to claims 18, 20, 22, and 23 are acknowledged.
Claims 1, 13, and 18-33 are pending.
Claims 18-33 are under examination on the merits.
Priority
This application 18/347,413 filed on 07/05/2023 is a divisional of U.S. Patent Application No. 16/758,726, filed on 04/23/2020 which is a 371 national phase application PCT/2018/059908 filed on 11/08/2018, and claims the benefit of provisional U.S. Patent Application No. 62/583,462, filed on 11/08/2017.
The priority date of claim18 and its dependent claims 19-33 is determined to be 11/08/2017, the filing date of provisional U.S. Patent Application No. 62/583,462.
Specification
The use of terms which are trade names or marks used in commerce (including New England Biolabs®), has been noted in this application. The term should be accompanied by the generic terminology; furthermore, the term should be capitalized wherever it appears or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM, or ® following the term.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Claim Interpretation
Claim 18 recites “a template strand having an identifier sequence, a first hairpin loop structure at a 5' region, and a second hairpin loop structure at a 3' region”. For purposes of examination a template strand is broadly interpreted as an oligonucleotide comprising the required structure.
Claim Rejections - 35 USC § 112(b)
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.
Claims 18-33 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.
Claim 18 recites the limitation “(a) providing a template strand having an identifier sequence -- wherein the identifier sequence is in a mid-region of the template strand --; (b) enzymatically extending the template strand from a 3' terminal end to meet the 5' terminal end such that the identifier sequence is double-stranded”. It is unclear that the identifier strand is not double-stranded before enzymatically extending the template strand.
Claims 19-33 are similarly indefinite because they directly or indirectly depend from claim 18.
Claim 19 recites the limitation “wherein the undesired products comprise molecules having the second hairpin loop structure prior to the cutting step and after the cutting step”. It is unclear what defines the undesired products. All molecules (duplex adapters) according to claim 18 have the second hairpin loop prior to the cutting step. For purposes of examination it is interpreted that the claim intends to require undesired products be molecules that retain a hairpin loop after the cutting step, specifically the cutting of the second cut site.
Claim 27 recites the limitation “the second cut comprises the modified nucleotide” There is insufficient antecedent basis for “the second cut” in the claim. Claim 18, which claim 27 depends from, recites “the second cut site”. There is also insufficient antecedent basis for “the modified nucleotide” in the claim. Claim 18, which claim 27 depends from, does not recite a modified nucleotide.
Claim 28 is similarly indefinite because it directly depends from claim 27.
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.
Claims 18-33 are rejected under 35 U.S.C. 103 as being unpatentable over Schmitt et al. (WO2013142389A1, on IDS dated 11/21/2023) in view of Hendrickson (US PGPub 20120238738, on IDS dated 11/21/2023), further in view of Schatz et al. (US PGPub 2006194202).
Regarding claim 18, Schmitt teaches Single Molecule Identifier adaptor synthesis. Regarding step (a), Schmitt teaches an adaptor molecule that includes a single molecule identifier (SMI) sequence comprising a degenerate or semi-degenerate DNA sequence (identifier sequence) and a ligation adaptor (para 9). The SMI adaptor may further comprise a hairpin shape (first hairpin loop structure) with a single-stranded nucleotide loop and a double-stranded stem portion, wherein the loop includes a cleavable linker (first cut site) (para 61) shown below:
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Regarding step (b), Schmitt teaches the complement of the degenerate lower arm sequence (N's) (identifier sequence) produced by polymerase extension (para 13, Fig. 2), which reads on extending the template strand from a 3' terminal end to meet the 5' terminal end such that the identifier sequence is double-stranded.
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Regarding step (c), Schmitt teaches cutting the hairpin uracil site, yielding an adaptor with a single stranded portion at a 5' end of the duplex adapter (para 62). Shown below:
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Further regarding step (a), Schmitt does not teach (i) a second hairpin loop structure at a 3' region, wherein the second hairpin loop structure comprises a second single-stranded nucleotide loop having a capture label and a 3' double-stranded stem portion having a second cut site.
Hendrickson teaches methods for making adapters, and teaches that loop adapters are convenient and easy to use, including adding the capacity to perform multiple enzymatic reactions (para 35).
Hendrickson teaches adapters with loops on both ends (first and second hairpin loops). The loop adapters may have one or more complementary (double-stranded) regions and a cleavage site may be located in a double-stranded (second cut site) or single-stranded (i.e. loop, first cut site) region of the adapter (para 9, Figs. 1A and 1B).
Hendrickson teaches that loop adapters provide multiple advantages including resistance to denaturation (para 57) and the addition of multiple enzymatic operations options (para 35 and 39-40).
Regarding the capture label, Schmitt teaches the SMI adaptor molecule, or a hairpin linker SMI adaptor molecule, may additionally contain modifications such as biotin (capture label)to facilitate affinity purification of target DNA that has ligated to the adaptor (para 66). Hendrickson also teaches a moiety such as biotin (capture label) may aid in separation, but does not teach a capture label in a single-stranded nucleotide loop.
Neither Schmitt nor Hendrickson teach a second single-stranded nucleotide loop having a capture label.
Schatz teaches a method for the manufacture of a nucleic acid molecule.
Schatz teaches providing a first at least partially double-stranded oligonucleotide which has a modification allowing the oligonucleotide to be coupled to a surface (a capture label) (abstract). Schatz teaches the modification is biotin (para 177, Fig. 1) located in a loop on one end of the nucleic acid, and further that a cleavage site is found in the double stranded region by the loop (Fig. 1).
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Further regarding step (a), Schmitt does not teach (ii) the identifier sequence is in a mid-region of the template strand between the 5' double-stranded stem portion (of the first hairpin loop structure) and the 3' double-stranded stem portion (of the second hairpin loop structure)
Hendrickson teaches loop adapters may additionally include sequence identifiers such as barcodes that are preferably located between the terminal region of the adapter and one or more modified components (e.g. between a first and second hairpin loop) (para 51). Hendrickson does not explicitly teach the identifier sequence is in a mid-region of the template strand between the two loops.
However, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Schmitt, Hendrickson, and Schatz to arrive at the instantly claimed invention as required for step (a). The modification would have entailed combining the adapter designs of Schatz and Hendrickson with Schmitt. Specifically, the modification would include adding a second loop adapter as in Schatz or Hendrickson, placing the identifier sequence between the loop adapters, and placing the capture label in a loop region of a terminal loop adapter as taught by Schatz. One would have been motivated by advantages offered by loop adapters and the ability to incorporate multiple enzymatic manipulations of an oligonucleotide strand as taught by Hendrickson. Determining the appropriate position to place an identifier sequence in an oligonucleotide strand or a capture label is considered a design choice that is part of routine optimization, which would be well within the purview of one skilled in the art. One would have been motivated to have the identifier sequence of Schmitt between the two loop adapters of Hendrickson in order to have the region accessible and allow enzymatic reactions such as cleavages to be isolated to the loop termini as taught by Hendrickson. One would further have been motivated to make the design choice of capture label placement by the need to place the capture label on an end of the oligonucleotide, for the purposes of purification as desired by both Schmitt and Hendrickson, and demonstrated for utility of nucleic acid synthesis by Schatz. There would have been a reasonable expectation of success given the underlying materials and methods are widely known, successfully demonstrated, and commonly used as evidenced by the prior art.
Further regarding step (c), Schmitt teaches an SMI adaptor may have a ligation adaptor (para 9), but does not teach the ligation adaptor is generated by cutting the second cut site. Schmitt does not teach (iii) cutting a second cut site to provide a duplex adapter having a ligation domain at a 3' end of the duplex adapter.
Hendrickson teaches a cleavage site may be located in a double-stranded (second cut site in a 3’ doble-stranded stem portion) but does not teach cutting the cleavage site in the stemloop provides a duplex adapter having a ligation domain at a 3' end of the duplex adapter.
Schatz teaches a cut site, Eam1104A, in a hairpin loop structure that provides a duplex having a ligation domain (Fig. 1).
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It would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Schmitt, Hendrickson, and Schatz to arrive at the instantly claimed invention of step (c). The modification would have entailed using the cut site of Hendrickson in the double stranded region of a terminal loop to generate the ligatable domain of Schmitt. Further, Schatz explicitly teaches the use of a cut site in the double-stranded region next to a loop to release a duplex comprising ligatable ends. Selecting the appropriate cut site to produce ligatable ends would have been a matter of judicious selection and routine optimization. One would have been motivated to use the advantages of targeting enzymatic operations to terminal loop adapters as taught by Hendrickson to produces the desired ligatable end of Schmitt. Determining the appropriate cut for producing ligatable ends would have been well within the purview of the skilled artisan. There would have been a reasonable expectation of success given the underlying materials and methods are widely known, successfully demonstrated, and commonly used as evidenced by the prior art.
Regarding claims 19 and 20, Schmitt teaches the SMI adaptor molecule, or a hairpin linker SMI adaptor molecule, may additionally contain modifications such as biotin (capture label) to facilitate affinity purification of target DNA that has ligated to the adaptor (para 66), which reads on separating the duplex adapter from undesired products. Hendrickson also teaches a moiety such as biotin (capture label) may aid in separation of products. However, neither Schmitt nor Hendrickson teach the undesired products comprise molecules having the second hairpin loop structure prior to the cutting step and after the cutting step.
Schatz teaches transferring elongated nucleotides (desired products) from a reaction (para 103 and Fig. 1). The elongated nucleic acids are double-stranded (duplex) (Fig. 1 below). Schatz further teaches the products that are not transferred (the undesired products) comprise oligonucleotides that have a second hairpin loop structure (comprising the capture label in the loop) (Fig. 1):
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It would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Schmitt and Hendrickson with Schmitt to arrive at the instantly claimed invention. The modification would have entailed using a cut site as taught by Hendrickson and Schatz to release a complete oligonucleotide after synthesis for use in downstream reactions as taught by Schatz. One would have been motivated by the ability to proceed with desired reactions with the finished oligonucleotide without the contamination of unprocessed oligonucleotides or adapters. There would have been a reasonable expectation of success given the underlying materials and methods are widely known, successfully demonstrated, and commonly used as evidenced by the prior art.
Regarding claim 21, Schmitt teaches the ligation adaptor includes a T-overhang, an A-overhang, a CG overhang, a blunt end, or any other ligatable sequence (para 30).
Regarding claim 22, Schmitt teaches the cleavable linker (first cut site) is uracil (U) (para 61), which reads on part (a), as shown below:
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Regarding claim 23, regarding the use of exonuclease, Schmitt teaches producing closed circles of captured material may help facilitate removal of non-captured DNA by exonuclease digestion given that circularized DNA will be protected from digestion by such enzymes (para 47). Hendrickson teaches unligated fragments and excess loop adapters can be removed by treating with one or more DNA exonucleases (para 12), and further, that the circular structure of an adapter-ligated double-stranded polynucleotide target also renders the construct resistant to exonuclease-degradation (para 60).
However, neither Schmitt, Hendrickson, not Schatz explicitly teach after the enzymatically extending step, the method further comprises applying an exonuclease.
It would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to use the teachings of Schmitt of Hendrickson to arrive at the instantly claimed invention. The modification would have entailed using the teachings of Hendrickson to use exonuclease to remove unligated fragments and excess loop adapters in the presence of the enzymatically extended oligonucleotides of Schmitt and Hendrickson. Enzymatically extending the complement of the identifier sequence of Schmitt in the presence of two loop adapters as taught by Hendrickson inherently results in a circular oligonucleotide strand. One of skill in the art would have been motivated to use the exonuclease cleanup step of Hendrickson with the modified, protected oligonucleotide adapters of Schmitt and Hendrickson. There would have been a reasonable expectation of success given the underlying materials and methods are widely known, successfully demonstrated, and commonly used as evidenced by the prior art.
Regarding claim 24, Schmitt teaches the hairpin shape (first hairpin loop structure) with a double-stranded stem portion includes complementary nucleotides (para 61), X and Y as shown below,
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which reads on part (a).
Regarding claim 25, Schmitt teaches the cleavable linker (first cut site) is uracil (U) (para 61).
Regarding claim 26, Schmitt teaches the cleavable linker uracil can be cleaved by uracil DNA glycosylase (para 61).
Regarding claim 27, Hendrickson teaches a loop adapter can be cleaved at or near a modified nucleotide (para 40), located in a double stranded region (stem-loop) (para 9) and that the modified nucleotide can be a ribonucleotide.
Regarding claim 28, Hendrickson teaches that when the modified nucleotide is a ribonucleotide, the adapter can be cleaved with an endoribonuclease (para 40). But does not teach the endoribonuclease is RNase H.
However, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Hendrickson to arrive at the instantly claimed invention. Determining an appropriate endoribonuclease would have been merely a matter of judicious selection and routine optimization which is well within the purview of the skilled artisan. There would have been a reasonable expectation of success given the underlying materials and methods are widely known, successfully demonstrated, and commonly used as evidenced by the prior art.
Regarding claims 29 and 30, Schatz teaches immobilizing a strand for nucleic acid synthesis (a template strand) via a capture label using a biotin group immobilized to a streptavidin coated surface (para 177).
Regarding claim 31, Schmitt teaches the adaptor contains a single molecule identifier (SMI) sequence (para 9).
Regarding claim 32, Schmitt teaches ligating the adaptor to a double-stranded target nucleic acid sequence (para 9).
Regarding claim 33, Schatz teaches the capture label on a hairpin loop is biotin (Fig. 1).
Conclusion
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/JESSICA GRAY/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682
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