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 I (claims 1-4, 6-7, 9-11, and 13-15) in the reply filed on 4/15/2026 is acknowledged.
Claim Status
Claims 1-4, 6-7, 9-11, 13-22, 24-25, 27-29, and 31-41 are pending.
Claims 16-22, 24-25, 27-29, and 31-41 withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 4/15/2026.
Claims 1-4, 6-7, 9-11, and 13-15 are being examined on the merits.
Information Disclosure Statement
The listing of references in the specification is not a proper information disclosure statement (see paragraphs [0029, 0060-0061, 0064, 0066-0067, 0084, 0088, 0091, 00116-00117, 00119, 00127, 00145]). 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be incorporated into the specification but must be submitted in a separate paper." Therefore, unless the references have been cited by the examiner on form PTO-892, they have not been considered.
Nucleotide and/or Amino Acid Sequence Disclosures
REQUIREMENTS FOR PATENT APPLICATIONS CONTAINING NUCLEOTIDE AND/OR AMINO ACID SEQUENCE DISCLOSURES
Items 1) and 2) provide general guidance related to requirements for sequence disclosures.
37 CFR 1.821(c) requires that patent applications which contain disclosures of nucleotide and/or amino acid sequences that fall within the definitions of 37 CFR 1.821(a) must contain a "Sequence Listing," as a separate part of the disclosure, which presents the nucleotide and/or amino acid sequences and associated information using the symbols and format in accordance with the requirements of 37 CFR 1.821 - 1.825. This "Sequence Listing" part of the disclosure may be submitted:
In accordance with 37 CFR 1.821(c)(1) via the USPTO patent electronic filing system (see Section I.1 of the Legal Framework for Patent Electronic System (https://www.uspto.gov/PatentLegalFramework), hereinafter "Legal Framework") as an ASCII text file, together with an incorporation-by-reference of the material in the ASCII text file in a separate paragraph of the specification as required by 37 CFR 1.823(b)(1) identifying:
the name of the ASCII text file;
ii) the date of creation; and
iii) the size of the ASCII text file in bytes;
In accordance with 37 CFR 1.821(c)(1) on read-only optical disc(s) as permitted by 37 CFR 1.52(e)(1)(ii), labeled according to 37 CFR 1.52(e)(5), with an incorporation-by-reference of the material in the ASCII text file according to 37 CFR 1.52(e)(8) and 37 CFR 1.823(b)(1) in a separate paragraph of the specification identifying:
the name of the ASCII text file;
the date of creation; and
the size of the ASCII text file in bytes;
In accordance with 37 CFR 1.821(c)(2) via the USPTO patent electronic filing system as a PDF file (not recommended); or
In accordance with 37 CFR 1.821(c)(3) on physical sheets of paper (not recommended).
When a “Sequence Listing” has been submitted as a PDF file as in 1(c) above (37 CFR 1.821(c)(2)) or on physical sheets of paper as in 1(d) above (37 CFR 1.821(c)(3)), 37 CFR 1.821(e)(1) requires a computer readable form (CRF) of the “Sequence Listing” in accordance with the requirements of 37 CFR 1.824.
If the "Sequence Listing" required by 37 CFR 1.821(c) is filed via the USPTO patent electronic filing system as a PDF, then 37 CFR 1.821(e)(1)(ii) or 1.821(e)(2)(ii) requires submission of a statement that the "Sequence Listing" content of the PDF copy and the CRF copy (the ASCII text file copy) are identical.
If the "Sequence Listing" required by 37 CFR 1.821(c) is filed on paper or read-only optical disc, then 37 CFR 1.821(e)(1)(ii) or 1.821(e)(2)(ii) requires submission of a statement that the "Sequence Listing" content of the paper or read-only optical disc copy and the CRF are identical.
Specific deficiencies and the required response to this Office Action are as follows:
Specific deficiency - This application fails to comply with the requirements of 37 CFR 1.821 - 1.825 because it does not contain a "Sequence Listing" as a separate part of the disclosure or a CRF of the “Sequence Listing.”. There are sequences present in Figure 10 of the provided Drawings, yet no Sequence Listing or CRF of the Sequence Listing is provided.
Required response - Applicant must provide:
A "Sequence Listing" part of the disclosure; together with
An amendment specifically directing its entry into the application in accordance with 37 CFR 1.825(a)(2);
A statement that the "Sequence Listing" includes no new matter as required by 37 CFR 1.821(a)(4); and
A statement that indicates support for the amendment in the application, as filed, as required by 37 CFR 1.825(a)(3).
If the "Sequence Listing" part of the disclosure is submitted according to item 1) a) or b) above, Applicant must also provide:
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3) and 1.125 inserting the required incorporation-by-reference paragraph, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter.
If the "Sequence Listing" part of the disclosure is submitted according to item 1) c) or d) above, applicant must also provide:
A CRF in accordance with 37 CFR 1.821(e)(1) or 1.821(e)(2) as required by 1.825(a)(5); and
A statement according to item 2) a) or b) above.
Specific deficiency – Nucleotide and/or amino acid sequences appearing in the drawings are not identified by sequence identifiers in accordance with 37 CFR 1.821(d). Sequence identifiers for nucleotide and/or amino acid sequences must appear either in the drawings or in the Brief Description of the Drawings.
Required response – Applicant must provide:
Replacement and annotated drawings in accordance with 37 CFR 1.121(d) inserting the required sequence identifiers;
AND/OR
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3) and 1.125 inserting the required sequence identifiers into the Brief Description of the Drawings, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter.
Specification
The disclosure is objected to because of the following informalities: In paragraphs [0012] and [0023] of the specification, the color “red” is referenced in regards to specific features of the drawings. However, color drawings are not provided. It is recommended to avoid using color to denote specific features of the drawings given that they are provided in black and white.
Appropriate correction is required.
The use of the terms “Life Technology”, “Oxford Nanopore”, and “Illumina” (paragraph [0071]) and “Qiagen”, “Ambion”, “Lucigen”, and “TRIzol” (paragraph [0085]), which are trade names or marks used in commerce, have been noted in this application. These terms should be accompanied by the generic terminology; furthermore the terms should be capitalized wherever they appear or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM, or ® following the terms.
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.
The examples above are not an exhaustive list of unmarked trade names or marks used in commerce throughout the specification. Please carefully read through and properly notate each instance.
Claim Objections
Claims 1-4 and 11 are objected to because of the following informalities:
Claim 1, line 8 reads “the PCR reaction comprises a plurality of fragments” and should read “the PCR reaction comprises a plurality of library fragments” to maintain consistent claim terminology (see line 4-5).
Claim 1, line 9 reads “dNTPS” and should read “[[dNTPS]]dNTPs”.
Claim 1, line 13 contains an unnecessary extra space after “(ii)”.
Claim 1, line 15 reads “a 3’-block that prevent polymerase extension” and should read “a 3’-block that prevents polymerase extension”.
Claim 1, line 17 reads “the template sequences” and should read “the double stranded template sequences” to maintain consistent claim terminology (see line 5).
Claim 1, line 18 reads “non-desired fragments, thereby bocking amplification of the non-desired fragments” and should read “[[non-desired]]non-desirable fragments, thereby bocking amplification of the [[non-desired]]non-desirable fragments” to maintain consistent claim terminology.
Claim 2 reads “wherein the one or more of the blocking oligonucleotides” and should read “wherein the one or more [[of the ]]blocking oligonucleotides”.
Claim 3 reads “when the polymerase has a 5’ to 3’ exonuclease activity, then the one or more of the blocking oligonucleotides comprise at the 5’ terminus, 1 to 5 nucleotides” and should read “when the polymerase has a 5’ to 3’ exonuclease activity, then the one or more [[of the ]]blocking oligonucleotides comprise at the 5’ terminus[[,]] 1 to 5 nucleotides”.
Claim 4 reads “when the polymerase has a 3’ to 5’ proofreading activity, then the one or more of the blocking oligonucleotides comprise at the 3’ terminus, 1 to 5 nucleotides” and should read “when the polymerase has a 3’ to 5’ proofreading activity, then the one or more [[of the ]]blocking oligonucleotides comprise at the 3’ terminus[[,]] 1 to 5 nucleotides”.
Claim 11 reads “template sequences from 18S rRNA, 5.8S rRNA, and/or 28S RNA” and should read “template sequences from 18S rRNA, 5.8S rRNA, and/or 28S rRNA”.
Appropriate correction is required.
Claim Interpretation
Claim 1 is directed to a method to “selectively deplete non-desirable fragments from amplified DNA or cDNA libraries by using one or more blocking oligonucleotides”. The only step of this method is performance of amplification on a plurality of library fragments that contain adapters in the presence of a blocking oligonucleotide. Because this achieves the depletion of non-desirable fragments in the creation of an amplified DNA or cDNA library, the starting material is interpreted to be a library of DNA or cDNA double stranded molecules that has not been amplified yet, but that has been processed to add adapters to the end or ends of said molecules.
Claim Rejections - 35 USC § 112b - Indefiniteness
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 1-4, 6-7, 9-11, and 13-15 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 1 recites the limitation "the one or more blocking primers" in line 17. There is insufficient antecedent basis for this limitation in the claim. For the purposes of examination, this is being interpretated to refer to the one or more blocking oligonucleotides as defined in lines 2-3. However, further clarification is required.
Claims 2-4, 6-7, 9-11, and 13-15 depend from claim 1, inherit these deficiencies, and are rejected on the same basis.
Claim 9 is directed to the method of claim 1 “wherein the adapter sequences are from Y-shaped adapters that have been ligated to each end of a template sequence”. However, a template sequence (“a double stranded template sequence including adapter sequences”) has already been defined in claim 1. Is this another type of template sequence that is present within the library which specifically has Y-shaped adapters or is this referring to the double stranded template sequences of the library as defined in claim 1? For the purposes of examination, “a template sequence” is interpreted to mean the double stranded template sequences of the library as defined in claim 1, prior to attachment of an adapter. However, further clarification is required.
Claim 10 is directed to the method of claim 1 “wherein the one or more blocking oligonucleotides bind to template sequences from mtDNA, rRNAs and/or globin”. The scope of “globin” is unclear. Is it meant that globin RNA is bound or that the genomic DNA sequence of globin is bound, or both? The scope of the claim is unclear and therefore indefinite. Clarification is required.
Claim Rejections - 35 USC § 102
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 the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1-4, 7, 9, and 13 are rejected under 35 U.S.C. 102(a)(1) and 102(a)(2) as being anticipated by Huang (Huang et al., WO 2017/142989 A1; cited on IDS of 9/29/2023).
Regarding claim 1: Huang teaches a method of selective amplification in which a DNA or cDNA library is amplified in the presence of a blocking oligonucleotide that prevents the amplification of an undesirable sequence (pg 2, ln 12-21, pg 2, ln 28, pg 11, ln 25-27, pg 12, ln 20-24). Huang teaches obtaining a plurality of double-stranded fragments that contain the desirable sequence and non-desirable sequences and adding adapters, and that said nucleic acid sample can be DNA or cDNA (pg 2, ln 12-15, pg 5, ln 18-20, pg 19, ln 20-21). Huang teaches performing PCR (pg 16, ln 6) on the target fragments with adapters with a forward and reverse primer in the presence of a blocker, a polymerase, and dNTPs (pg 15, ln 29-30). While Huang does not explicitly list dNTPs in the reaction, dNTPs are evidenced by the successful creation of amplified product that could be sequenced (pg 44, ln 31 and pg 45, ln 1-3). Additionally, Huang teaches that PCR amplification is performed in the presence of “essential ingredients” (pg 10, ln 21) which those of skill in the art know to include dNTPs. Huang teaches of blockers that comprise a 3’ block that prevents polymerase extension on the 3’ terminus of the oligonucleotide (“non-extendable” blocker, pg 12, ln 20-24) and which contain 3’ or 5’ phosphorothioate backbone modifications to protect against 3’ or 5’ exonuclease activity (pg 12, ln 1-5). Huang teaches that the blocking oligonucleotides bind to the sequences of non-desirable templates and prevents amplification of those fragments (pg 15, ln 30-31, pg 16, ln 1, pg 18, ln 29-32).
Regarding claim 2: Huang teaches that the one or more blocking oligonucleotides are less than 100 nt in length with a lower limit of 5 nt (significantly overlaps with 15 nt to 100 nt; pg 11, ln 27-30).
Regarding claims 3 and 4: Huang teaches that the blocking oligonucleotides contain 3’ or 5’ phosphorothioate backbone modifications to protect against 3’ or 5’ exonuclease activity (pg 12, ln 1-5).
Regarding claim 7: Huang teaches that the amplified libraries comprises sequences from cDNA or from gDNA (pg 5, ln 27-30, pg 19, ln 20-21).
Regarding claim 9: Huang teaches the use of Y-shaped adapters ligated at each end of a template sequence (pg 8, ln 17, pg 9, ln 14).
Regarding claim 13: Huang teaches that the libraries can be analyzed by subsequence NGS sequencing (pg 18, line 27-28).
Claim Rejections - 35 USC § 103
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.
Claim 6 is rejected under 35 U.S.C. 103 as being unpatentable over Huang (Huang et al., WO 2017/142989 A1; cited on IDS of 9/29/2023) in view of Vestheim (Vestheim et al., PCR Protocols 2010).
The teachings of Huang as they apply to claim 1, from which claim 6 depends, are detailed above. Relevant to the instantly rejected claim, Huang teaches a method of selectively amplifying a DNA or cDNA library in which a plurality of fragments with adapters are amplified in the presence of a non-extendable blocker oligonucleotide. Huang teaches that this blocker oligonucleotide contains a 3’ blocking moiety that prevents extension.
Huang does not teach what the 3’ blocking moiety is that prevents extension by a polymerase. However, 3’ modification to blocking oligonucleotides that prevents extension by a polymerase in a PCR amplification reaction are known in the art, as taught by Vestheim.
Vestheim teaches about blocking oligonucleotides that bind to unwanted DNA templates to prevent their amplification in favor of amplification of wanted or rarer sequences (Abstract). Vestheim teaches that common 3’ blocking moieties include a C3 spacer, 3’ phosphate group, and 3’ inverted end base (2.1.1 Standard DNA Oligos Modified to Not Prime Amplification).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Huang to include the specific 3’ blocking moieties as taught by Vestheim. One would be motivated to use said blocking moieties given the assertion by Vestheim that these are standard oligonucleotides used in the art to prevent priming by a polymerase and are modification that are “offered from most suppliers of custom oligonucleotides” (2.1.1 Standard DNA Oligos Modified to Not Prime Amplification). One would have a reasonable expectation of success given that Vestheim teaches that these types of oligos with 3’ modification are of standard use in the art of selective amplification.
Claims 10 and 11 are rejected under 35 U.S.C. 103 as being unpatentable over Huang (Huang et al., WO 2017/142989 A1; cited on IDS of 9/29/2023) in view of Shaffer (Shaffer et al., WO 2020/068559 A1).
The teachings of Huang as they apply to claim 1, from which claims 10 and 11 depend, are detailed above. Relevant to the instantly rejected claim, Huang teaches a method of selectively amplifying a DNA or cDNA library in which a plurality of fragments with adapters are amplified in the presence of a non-extendable blocker oligonucleotide.
Huang does not teach that a target of the blocker oligonucleotides (or the non-desirable sequence) is from rRNA, more specifically 18S rRNA, 5.8S rRNA, and/or 28S rRNA. However, depletion of rRNA (18S, 5.8S, and 28S) from an amplified sequencing library by blocking oligonucleotides is known in the art, as taught by Shaffer.
Shaffer teaches depletion of unwanted RNA species from RNA samples in the construction of a sequencing library (pg 5, ln 13-17). Shaffer teaches designing blocking oligonucleotides against unwanted RNA species such as 28S rRNA, 18S rRNA, and 5.8S rRNA (pg 8, ln 20-22). Shaffer teaches that preferably multiple rRNA types are depleted using multiple blocking oligonucleotides (pg 8, ln 23-25 and pg 3, ln 28-29, pg 4, ln 1-3).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Huang to specifically target unwanted rRNA species as taught by Shaffer. One would be motivated to do so given the assertion by Shaffer that the majority of transcriptome sequencing budget is taken up by unwanted species of RNA (such as rRNA, mtrRNA, and globin mRNA) that can comprise as much as 80% of the rRNA sample, which has motivated the creation of methods of depletion of said species from NGS libraries (pg 1, ln 15-21). While Shaffer specifically teaches using non-extendable blocking oligonucleotides for the prevention of cDNA synthesis of these unwanted species, the same principles of blocking oligonucleotide design and hybridization to prevent amplification of cDNA would be obvious to those skilled in the art aiming to deplete these unwanted species from cDNA libraries, according to the methodology of Huang. One would have a reasonable expectation of success given that Shaffer successfully depletes unwanted rRNA from NGS libraries using blocking oligonucleotides complementary to target sequences.
Claims 14 and 15 are rejected under 35 U.S.C. 103 as being unpatentable over Huang (Huang et al., WO 2017/142989 A1; cited on IDS of 9/29/2023) in view of van Dijk (van Dijk et al., Experimental Cell Research 2014; cited on IDS of 9/29/2023).
The teachings of Huang as they apply to claim 1, from which claims 10 and 11 depend, are detailed above. Relevant to the instantly rejected claim, Huang teaches a method of selectively amplifying a DNA or cDNA library in which a plurality of fragments with adapters are amplified in the presence of a non-extendable blocker oligonucleotide. Huang teaches that RNA samples, in the creation of a cDNA library, can be sheared (fragmented) and then reverse transcribed (pg 21, ln 17-20).
Regarding claim 14: While Huang does teach blunt-ending and ligation of adapters to sheared ends double-stranded templates, Huang does not teach the specific methodology in which the cDNA created from reverse transcribing the RNA fragments undergoes blunt ending, addition of an A nucleotide to the 3’ end of the blunt ended cDNA, or ligating the A-tailed cDNA with adapters comprising a non-complemented T nucleotide at the 3’ end. However, this methodology of cDNA library adapter attachment is known in the art, as taught by van Dijk.
Van Dijk teaches a common RNA-seq preparation method of end repairing double-stranded cDNA, A-tailing the end-repaired double-stranded cDNA, and then ligating adapters to both ends of said A-tailed dsDNA using adapters with a non-complemented T nucleotide at the 3’ end (Figure 2A).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Huang to include the library preparation steps as taught by van Dijk for attachment of the adapters. One would be motivated to do so given the assertion by van Dijk that this methodology is one of the most common methods of library preparation for RNA samples (Figure 2 legend). One would have a reasonable expectation of success given that van Dijk teaches that many current library preparation kits follow the RNA ligation method (Figure 2 legend and Summary and practical considerations).
Regarding claim 15: van Dijk teaches prior to reverse transcribing RNA fragments into cDNA, the RNA sample is treated to deplete rRNA sequences from the RNA sample (RNA-sequencing).
Conclusion
No claims are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to KAILEY E CASH whose telephone number is (571)272-0971. The examiner can normally be reached Monday-Friday 8:30am-6pm ET.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Anne Gussow can be reached at (571)272-6047. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/KAILEY ELIZABETH CASH/Examiner, Art Unit 1683
/ANNE M. GUSSOW/Supervisory Patent Examiner, Art Unit 1683
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