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 .
Status of the Claims
Claims 1-14 are currently pending and under exam herein.
Claims 1-14 are rejected.
Priority
The instant application is a 371 of PCT KR2021/018746 filed on 12/10/2021 which claims priority from KR10-2020-0173648 filed on 12/11/2020. Thus, the effective filing date of the instant application is 12/11/2020.
Drawings
The Drawings filed on 05/24/2023 were considered.
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 05/24/2023 is in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statement has been considered by the examiner.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 1-14 are rejected under 35 U.S.C. 101 because the claimed invention is directed to an abstract idea without significantly more. The claims recite: (a) mathematical concepts, (e.g., mathematical relationships, formulas or equations, mathematical calculations); and (b) mental processes, i.e., concepts performed in the human mind, (e.g., observation, evaluation, judgement, opinion).
Subject matter eligibility evaluation in accordance with MPEP 2106:
Eligibility Step 1: Claims 1-14 are directed to a method to design an oligonucleotide of interest.
[Step 1: YES]
Eligibility Step 2A: First it is determined in Prong One whether a claim recites a judicial exception, and if
so, then it is determined in Prong Two whether the recited judicial exception is integrated into a
practical application of that exception.
Eligibility Step 2A Prong One: In determining whether a claim is directed to a judicial exception,
examination is performed that analyzes whether the claim recites a judicial exception, i.e., whether a
law of nature, natural phenomenon, or abstract idea is set forth or described in the claim.
Independent claim 1 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
(a) inputting information about a wild-type target nucleic acid sequence and a nucleotide mutation of interest; wherein the wild-type target nucleic acid sequence is a target nucleic acid sequence not comprising the nucleotide mutation of interest, the information includes information about (i) the wild-type target nucleic acid sequence, (ii) a position of the nucleotide mutation of interest occurring in the wild-type target nucleic acid sequence and (iii) wild-type and mutant bases at the position of the nucleotide mutation of interest, and wherein the position of the nucleotide mutation of interest is expressed as a start position and an end position, (mental process and/or mathematical concept)
(b) providing the wild-type target nucleic acid sequence and a mutant target nucleic acid sequence by using the input information; wherein the mutant target nucleic acid sequence is a target nucleic acid sequence comprising the nucleotide mutation of interest, and the wild-type and mutant target nucleic acid sequences include forward and reverse wild-type and mutant target nucleic acid sequences, respectively, (mental process)
(c) providing a first oligonucleotide candidate group for the mutant target nucleic acid sequence by designing oligonucleotides used to detect the nucleotide mutation of interest in a predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence; (mental process and/or mathematical concept)
(d) providing oligonucleotides satisfying the following selection criteria as a second oligonucleotide candidate group for the mutant target nucleic acid sequence by analyzing matching of the wild-type target nucleic acid sequence with the oligonucleotides included in the first oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the selection criteria include that (i) the number of mismatches between the wild-type target nucleic acid sequence and a predetermined region at the 5′-end, middle or 3′-end of an oligonucleotide is one or more; or (ii) the ratio of mismatches between the wild-type target nucleic acid sequence and an oligonucleotide is a predetermined value or more; and (mental process and/or mathematical concept)
(e) providing a third oligonucleotide candidate group by selecting oligonucleotides from the second oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the third oligonucleotide candidate group is used to detect the nucleotide mutation of interest in the target nucleic acid sequence. (mental process)
Dependent claim 2 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the nucleotide mutation of interest is a substitution, an inversion, an insertion, a deletion, a duplication, or a combination thereof. (mental process)
Dependent claim 3 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the oligonucleotide is a probe and/or a primer. (mental process)
Dependent claim 4 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the wild-type target nucleic acid sequence in step (a) has a predetermined length comprising a position of the nucleotide mutation of interest. (mental process and/or mathematical concept)
Dependent claim 5 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the oligonucleotides in step (c) are designed to have matching or complementary sequences to the predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence. (mental process and/or mathematical concept)
Dependent claim 6 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the oligonucleotides in step (c) are designed according to lengths at positions from a predetermined position upstream of the start position of the nucleotide mutation of interest within the mutant target nucleic acid sequence to the end position thereof. (mental process and/or mathematical concept)
Dependent claim 7 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the oligonucleotides in step (c) are designed to satisfy at least one of the following conditions: (i) a length of 10-60 nucleotides; (ii) a Tm value of 50-85° C.; (iii) exclusion of a G-run sequence with at least three Gs; and (iv) a GC content of 40% or more in the 5′-end portion. (mental process and/or mathematical concept)
Dependent claim 8 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the mismatches (i) in step (d) are minimum mismatches between the wild-type target nucleic acid sequence and the predetermined region of the 5′-end, middle, or 3′-end of the oligonucleotide, and the mismatches (ii) in step (d) are minimum mismatches between the wild-type target nucleic acid sequence and the oligonucleotide. (mental process and/or mathematical concept)
Dependent claim 9 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the method further comprises, after step (d), d-1) arranging the oligonucleotides included in the second oligonucleotide candidate group by giving ranks according to at least one of the following arrangement criteria: (mental process and/or mathematical concept)
(i) the position of a mismatch in an oligonucleotide as a result of the matching analysis in step (d); wherein the closer the position of the mismatch is to the 5′-end, 3′-end, or middle, the higher the rank, (mental process and/or mathematical concept)
(ii) the number of mismatches of an oligonucleotide as a result of the matching analysis in step (d); wherein the larger the number of the mismatches, the higher the rank, (mental process and/or mathematical concept)
(iii) a Tm value of an oligonucleotide; wherein the higher the Tm value, the higher the rank, (mental process and/or mathematical concept)
(iv) a GC content in a predetermined region of the 5′-end, middle, or 3′-end of an oligonucleotide; wherein the higher the GC content, the higher the rank, (mental process and/or mathematical concept)
(v) the number of consecutive G bases included in an oligonucleotide; wherein the smaller the number of consecutive G bases, the higher the rank, (mental process and/or mathematical concept)
(vi) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when the oligonucleotide forms the homodimer; wherein the smaller the number or proportion, the higher the rank, (mental process and/or mathematical concept)
(vii) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, and (mental process and/or mathematical concept)
(viii) a length; wherein the shorter the length, the higher the rank. (mental process and/or mathematical concept)
Dependent claim 10 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the third oligonucleotide candidate group in step (e) is selected by a method comprising the following steps: (mental process and/or mathematical concept)
(e-1) arranging by giving ranks according to at least one of the following arrangement criteria to oligonucleotides having the same start position for designing an oligonucleotide among the oligonucleotides included in the second oligonucleotide candidate group; and (mental process and/or mathematical concept)
(e-2) selecting the highest ranked oligonucleotide from oligonucleotides having the same start position for designing the arranged oligonucleotides, and wherein the arrangement criteria include the following: (mental process and/or mathematical concept)
(i) the position of a mismatch in an oligonucleotide as a result of the matching analysis in step (d); wherein the closer the position of the mismatch is to the 5′-end, 3′-end, or middle, the higher the rank, (mental process and/or mathematical concept)
(ii) the number of mismatches in an oligonucleotide as a result of the matching analysis in step (d); wherein the larger the number of the mismatches, the higher the rank, (mental process and/or mathematical concept)
(iii) a Tm value of an oligonucleotide; wherein the higher the Tm value, the higher the rank, (mental process and/or mathematical concept)
(iv) a GC content in a predetermined region of the 5′-end, middle, or 3′-end of an oligonucleotide; wherein the higher the GC content, the higher the rank, (mental process and/or mathematical concept)
(v) the number of consecutive G bases included in an oligonucleotide; wherein the smaller the number of consecutive G bases, the higher the rank, (mental process and/or mathematical concept)
(vi) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when the oligonucleotide forms the homodimer; wherein the smaller the number or proportion, the higher the rank, (mental process and/or mathematical concept)
(vii) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, and (mental process and/or mathematical concept)
(viii) a length; wherein the shorter the length, the higher the rank. (mental process and/or mathematical concept)
Dependent claim 11 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the first to third oligonucleotide candidate groups are first to third probe candidate groups, respectively, and the method further comprises, after step (e), the following steps: (f) providing a primer candidate group for the mutant target nucleic acid sequence by designing primers to amplify a predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence; and (mental process)
(g) providing a combination of a probe and primers by combining the third probe candidate group and the primer candidate group. (mental process)
Dependent claim 12 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the primers in step (f) are designed to satisfy at least one of the following conditions:
(i) a Tm value of 40-70° C.; (mental process and/or mathematical concept)
(ii) a length of 15-50 bp nucleotides; and (mental process and/or mathematical concept)
(iii) exclusion of a G-run sequence with at least five Gs. (mental process and/or mathematical concept)
Dependent claim 13 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
wherein the method further comprises, after step (f), the following steps: (mental process and/or mathematical concept)
(f-1) arranging by giving ranks according to at least one of the following arrangement criteria to primers having the same start position for designing a primer among the primers included in the primer candidate group; and (mental process and/or mathematical concept)
(f-2) selecting the highest ranked primer from primers having the same start position for designing the arranged primers, and wherein the arrangement criteria include the following: (mental process and/or mathematical concept)
(i) the number or proportion of use of a degenerate base and/or universal base introduced into a primer; wherein the smaller the number or proportion of use, the higher the rank, (mental process and/or mathematical concept)
(ii) the number of primer patterns generated by the introduction of a degenerate base; wherein the smaller the number of patterns, the higher the rank, (mental process and/or mathematical concept)
(iii) the number of (A)n, (T)n, or (C)n mononucleotide run sequences; wherein the smaller the number of the sequences, the higher the rank, (mental process and/or mathematical concept)
(iv) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when a primer forms the homodimer; wherein the smaller the number or proportion, the higher the rank, (mental process and/or mathematical concept)
(v) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, (mental process and/or mathematical concept)
(vi) a Tm value; wherein the higher the Tm value, the higher the rank, (mental process and/or mathematical concept)
(vii) a GC content; wherein the more the GC content, the higher the rank, and (mental process and/or mathematical concept)
(viii) a length; wherein the shorter the length, the higher the rank (mental process and/or mathematical concept)
Independent claim 14 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
(a) inputting information about a wild-type target nucleic acid sequence and a nucleotide mutation of interest; wherein the wild-type target nucleic acid sequence is a target nucleic acid sequence not comprising the nucleotide mutation of interest, the information includes information about (i) the wild-type target nucleic acid sequence, (ii) a position of the nucleotide mutation of interest occurring in the wild-type target nucleic acid sequence and (iii) wild-type and mutant bases at the position of the nucleotide mutation of interest, and wherein the position of the nucleotide mutation of interest is expressed as a start position and an end position, (mental process)
(b) providing the wild-type target nucleic acid sequence and a mutant target nucleic acid sequence by using the input information; wherein the mutant target nucleic acid sequence is a target nucleic acid sequence comprising the nucleotide mutation of interest, and the wild-type and mutant target nucleic acid sequences include forward and reverse wild-type and mutant target nucleic acid sequences, respectively, (mental process)
(c) providing a first oligonucleotide candidate group for the mutant target nucleic acid sequence by designing oligonucleotides used to detect the nucleotide mutation of interest in a predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence; (mental process)
(d) providing oligonucleotides satisfying the following selection criteria as a second oligonucleotide candidate group for the mutant target nucleic acid sequence by analyzing matching of the wild-type target nucleic acid sequence with the oligonucleotides included in the first oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the selection criteria include that (i) the number of mismatches between the wild-type target nucleic acid sequence and a predetermined region at the 5′-end, middle or 3′-end of an oligonucleotide is one or more; or (ii) the ratio of mismatches between the wild-type target nucleic acid sequence and an oligonucleotide is a predetermined value or more; and (mental process)
(e) providing a third oligonucleotide candidate group by selecting oligonucleotides from the second oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the third oligonucleotide candidate group is used to detect the nucleotide mutation of interest in the target nucleic acid sequence. (mental process)
The MPEP states the following “Accordingly, the "mental processes" abstract idea grouping is defined as concepts performed in the human mind, and examples of mental processes include observations, evaluations, judgments, and opinions. A discussion of concepts performed in the human mind, as well as concepts that cannot practically be performed in the human mind and thus are not "mental processes", is provided below with respect to point A. The courts do not distinguish between mental processes that are performed entirely in the human mind and mental processes that require a human to use a physical aid (e.g., pen and paper or a slide rule) to perform the claim limitation. See, e.g., Benson, 409 U.S. at 67, 65, 175 USPQ at 674-75, 674 (noting that the claimed "conversion of [binary-coded decimal] numerals to pure binary numerals can be done mentally," i.e., "as a person would do it by head and hand."); Synopsys, Inc. v. Mentor Graphics Corp., 839 F.3d 1138, 1139, 120 USPQ2d 1473, 1474 (Fed. Cir. 2016) (holding that claims to a mental process of "translating a functional description of a logic circuit into a hardware component description of the logic circuit" are directed to an abstract idea, because the claims "read on an individual performing the claimed steps mentally or with pencil and paper"). Mental processes performed by humans with the assistance of physical aids such as pens or paper are explained further below with respect to point B. Nor do the courts distinguish between claims that recite mental processes performed by humans and claims that recite mental processes performed on a computer. As the Federal Circuit has explained, "[c]ourts have examined claims that required the use of a computer and still found that the underlying, patent-ineligible invention could be performed via pen and paper or in a person’s mind." Versata Dev. Group v. SAP Am., Inc., 793 F.3d 1306, 1335, 115 USPQ2d 1681, 1702 (Fed. Cir. 2015). See also Intellectual Ventures I LLC v. Symantec Corp., 838 F.3d 1307, 1318, 120 USPQ2d 1353, 1360 (Fed. Cir. 2016) (‘‘[W]ith the exception of generic computer-implemented steps, there is nothing in the claims themselves that foreclose them from being performed by a human, mentally or with pen and paper.’’); Mortgage Grader, Inc. v. First Choice Loan Servs. Inc., 811 F.3d 1314, 1324, 117 USPQ2d 1693, 1699 (Fed. Cir. 2016) (holding that computer-implemented method for "anonymous loan shopping" was an abstract idea because it could be "performed by humans without a computer"). Mental processes recited in claims that require computers are explained further below with respect to point C. Because both product and process claims may recite a "mental process", the phrase "mental processes" should be understood as referring to the type of abstract idea, and not to the statutory category of the claim. The courts have identified numerous product claims as reciting mental process-type abstract ideas, for instance the product claims to computer systems and computer-readable media in Versata Dev. Group. v. SAP Am., Inc., 793 F.3d 1306, 115 USPQ2d 1681 (Fed. Cir. 2015). This concept is explained further below with respect to point D. claim to "collecting information, analyzing it, and displaying certain results of the collection and analysis," where the data analysis steps are recited at a high level of generality such that they could practically be performed in the human mind, Electric Power Group v. Alstom, S.A., 830 F.3d 1350, 1353-54, 119 USPQ2d 1739, 1741-42, claims to "comparing BRCA sequences and determining the existence of alterations," where the claims cover any way of comparing BRCA sequences such that the comparison steps can practically be performed in the human mind, University of Utah Research Foundation v. Ambry Genetics, 774 F.3d 755, 763, 113 USPQ2d 1241, 1246 (Fed. Cir. 2014); a claim to collecting and comparing known information (claim 1), which are steps that can be practically performed in the human mind, Classen Immunotherapies, Inc. v. Biogen IDEC, 659 F.3d 1057, 1067, 100 USPQ2d 1492, 1500 (Fed. Cir. 2011)”
The abstract ideas recited in the claims are evaluated under the broadest reasonable interpretation (BRI) of the claim limitations when read in light of and consistent with the specification. As noted in the foregoing section, the claims are determined to contain limitations that can practically be performed in the human mind with the aid of a pencil and paper, and therefore recite judicial exceptions from the mental process grouping of abstract ideas. Additionally, the recited limitations that are identified as judicial exceptions from the mathematical concepts grouping of abstract ideas are abstract ideas irrespective of whether or not the limitations are practical to perform in the human mind.
Therefore, claims 1-14 recite an abstract idea as the dependent claims will inherit the abstract ideas from the independent claims.
[Step 2A Prong One: YES]
Eligibility Step 2A Prong Two: In determining whether a claim is directed to a judicial exception, further
examination is performed that analyzes if the claim recites additional elements that when examined as a
whole integrates the judicial exception(s) into a practical application (MPEP 2106.04(d)). A claim that
integrates a judicial exception into a practical application will apply, rely on, or use the judicial exception
in a manner that imposes a meaningful limit on the judicial exception. The claimed additional elements
are analyzed to determine if the abstract idea is integrated into a practical application (MPEP
2106.04(d)(I); MPEP 2106.05(a-h)). If the claim contains no additional elements beyond the abstract
idea, the claim fails to integrate the abstract idea into a practical application (MPEP 2106.04(d)(III)).
The judicial exceptions identified in Eligibility Step 2A Prong One are not integrated into a practical application because of the reasons noted below.
The additional element in independent claim 1 includes:
A computer-implemented method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, comprising:
The additional element in independent claim 14 includes:
A computer readable storage medium containing instructions to configure a processor to perform a method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, the method comprising:
The additional elements of a computer-implemented method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, comprising (Claim 1), a computer readable storage medium containing instructions to configure a processor to perform a method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, the method comprising (Claim 14) fail to integrate a judicial exception into a practical application merely reciting the words "apply it" (or an equivalent) with the judicial exception, or merely including instructions to implement an abstract idea on a computer, or merely using a computer as a tool to perform an abstract idea, as discussed in MPEP § 2106.05(f).
Thus, the additionally recited elements merely invoke a computer as a tool, and/or amount to insignificant extra-solution data gathering activity, and as such, when all limitations in claims 1-14 have been considered as a whole, the claims are deemed to not recite any additional elements that would integrate a judicial exception into a practical application, and therefore claims 1-14 are directed to an abstract idea (MPEP 2106.04(d)).
[Step 2A Prong Two: NO]
Eligibility Step 2B: Because the claims recite an abstract idea, and do not integrate that abstract idea into a practical application, the claims are probed for a specific inventive concept. The judicial exception alone cannot provide that inventive concept or practical application (MPEP 2106.05). Identifying whether the additional elements beyond the abstract idea amount to such an inventive concept requires considering the additional elements individually and in combination to determine if they amount to significantly more than the judicial exception (MPEP 2106.05A i-vi).
The claims do not include any additional elements that are sufficient to amount to significantly more than the judicial exception(s) because of the reasons noted below.
The additional elements recited in claims 1-14 are identified above, and carried over from Step 2A: Prong Two along with their conclusions for analysis at Step 2B. Any additional element or combination of elements that was considered to be insignificant extra-solution activity at Step 2A: Prong Two was re-evaluated at Step 2B, because if such re-evaluation finds that the element is unconventional or otherwise more than what is well-understood, routine, conventional activity in the field, this finding may indicate that the additional element is no longer considered to be insignificant; and all additional elements and combination of elements were evaluated to determine whether any additional elements or combination of elements are other than what is well-understood, routine, conventional activity in the field, or simply append well-understood, routine, conventional activities previously known to the industry, specified at a high level of generality, to the judicial exception, per MPEP 2106.05(d).
The additional elements of a computer-implemented method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, comprising (Claim 1), a computer readable storage medium containing instructions to configure a processor to perform a method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, the method comprising (Claim 14) are conventional fail to integrate a judicial exception into a practical application merely reciting the words "apply it" (or an equivalent) with the judicial exception, or merely including instructions to implement an abstract idea on a computer, or merely using a computer as a tool to perform an abstract idea, as discussed in MPEP § 2106.05(f).
When taken alone, all additional elements in claims 1-14 do not amount to significantly more than the above-identified judicial exception(s). Even when evaluated as a combination, the additional elements fail to transform the exception(s) into a patent-eligible application of that exception. Thus, claims 1-14 are deemed to not contribute an inventive concept, i.e., amount to significantly more than the judicial exception(s) (MPEP 2106.05(II)).
[Step 2B: NO]
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 1-14 are rejected under 35 U.S.C. 103 as being unpatentable over Wangkumhang et al. (Wangkumhang, P.; Chaichoompu, K.; Ngamphiw, C.; Ruangrit, U.; Chanprasert, J.; Assawamakin, A.; Tongsima, S. WASP: A Web-Based Allele-Specific PCR Assay Designing Tool for Detecting SNPs and Mutations. BMC Genomics 2007, 8 (1)) in further view of Untergasser et al. (Untergasser, A.; Cutcutache, I.; Koressaar, T.; Ye, J.; Faircloth, B. C.; Remm, M.; Rozen, S. G. Primer3—New Capabilities and Interfaces. Nucleic Acids Research 2012, 40 (15), e115–e115.) in further view of You et al. (You, F. M.; Huo, N.; Gu, Y. Q.; Luo, M.-C.; Ma, Y.; Hane, D.; Lazo, G. R.; Dvorak, J.; Anderson, O. D. BatchPrimer3: A High Throughput Web Application for PCR and Sequencing Primer Design. BMC bioinformatics 2008, 9, 253) in further view of Ye et al. (Ye, J.; Coulouris, G.; Zaretskaya, I.; Cutcutache, I.; Rozen, S.; Madden, T. L. Primer-BLAST: A Tool to Design Target-Specific Primers for Polymerase Chain Reaction. BMC Bioinformatics 2012, 13 (1).) in further view of Rose et al. (Rose, T. M.; Schultz, E. R.; Henikoff, J. G.; Pietrokovski, S.; McCallum, C. M.; Henikoff, S. Consensus-Degenerate Hybrid Oligonucleotide Primers for Amplification of Distantly Related Sequences. Nucleic Acids Research 1998, 26 (7), 1628–1635.). The italicized text corresponds to the instant claim limitations.
With respect to the limitations of Claims 1, 2, 3, 4, 11, 14, Wangkumhang et al. teaches WASP offers a tool for designing AS primers for both SNPs and mutations (which include inversions of nucleotides). By integrating the database for known SNPs (using gene ID or rs number), this tool facilitates the awkward process of getting flanking sequences and other related information from public SNP databases. It takes into account the underlying destabilizing effect to ensure the effectiveness of designed primers. With user-friendly SVG interface, WASP intuitively presents resulting designed primers (Conclusion) both wild and mutant type templates are generated separately and the second input format is constructed to facilitate the AS primer design for novel SNPs and/or mutations. The target variations and their corresponding 5' and 3' flanking sequences must be entered to the given text box or upload a file containing the information to the server. Instead of constructing SQL queries described previously, the input data will be converted to the internal forms by the parsing module and later sent to the AS primer analysis module. For example, Figure 2 demonstrates two possible sample inputs that WASP would except to calculate the corresponding AS primer(s). The input format for this section comprises 5' and 3' flanking sequences or a target variation, which is described in IUPAC (Figure 2a) or bracketing format (Figure 2b). Note that for both inputs (querying from database and manually entering), their primer conditions can be modified such as primer length, melting temperature (Tm), GC content for primer oligos and other standard primer design parameters (1) Input module, 2nd paragraph, A computer-implemented method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, comprising: (Claim 1), A computer readable storage medium containing instructions to configure a processor to perform a method for preparing an oligonucleotide used to detect a nucleotide mutation of interest in a target nucleic acid sequence, the method comprising (Claim 14), (a) inputting information about a wild-type target nucleic acid sequence and a nucleotide mutation of interest; wherein the wild-type target nucleic acid sequence is a target nucleic acid sequence not comprising the nucleotide mutation of interest (Claim 1, Claim 14) (b) providing the wild-type target nucleic acid sequence and a mutant target nucleic acid sequence by using the input information; wherein the mutant target nucleic acid sequence is a target nucleic acid sequence comprising the nucleotide mutation of interest (Claim 1, Claim 14) (e) providing a third oligonucleotide candidate group by selecting oligonucleotides from the second oligonucleotide candidate group for the mutant target nucleic acid sequence (Claim 1, Claim 14); wherein the third oligonucleotide candidate group is used to detect the nucleotide mutation of interest in the target nucleic acid sequence. (Claim 2) wherein the oligonucleotide is a probe and/or a primer. (Claim 3) wherein the wild-type target nucleic acid sequence in step (a) has a predetermined length comprising a position of the nucleotide mutation of interest (Claim 4), (g) providing a combination of a probe and primers by combining the third probe candidate group and the primer candidate group. (Claim 11)
Wangkumhang et al. does not explicitly teach
(a) inputting information about a wild-type target nucleic acid sequence and a nucleotide mutation of interest; wherein the wild-type target nucleic acid sequence is a target nucleic acid sequence not comprising the nucleotide mutation of interest, the information includes information about (i) the wild-type target nucleic acid sequence, (ii) a position of the nucleotide mutation of interest occurring in the wild-type target nucleic acid sequence and (iii) wild-type and mutant bases at the position of the nucleotide mutation of interest (Claim 1, Claim 14)
and the wild-type and mutant target nucleic acid sequences include forward and reverse wild-type and mutant target nucleic acid sequences, respectively,(Claim 1, Claim 14),
(c) providing a first oligonucleotide candidate group for the mutant target nucleic acid sequence by designing oligonucleotides used to detect the nucleotide mutation of interest in a predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence; (Claim 1, Claim 14)
wherein the oligonucleotides in step (c) are designed to have matching or complementary sequences to the predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence. (Claim 5)
wherein the oligonucleotides in step (c) are designed according to lengths at positions from a predetermined position upstream of the start position of the nucleotide mutation of interest within the mutant target nucleic acid sequence to the end position thereof (Claim 6)
wherein the oligonucleotides in step (c) are designed to satisfy at least one of the following conditions: (i) a length of 10-60 nucleotides; (ii) a Tm value of 50-85° C.; (iii) exclusion of a G-run sequence with at least three Gs; and (iv) a GC content of 40% or more in the 5′-end portion (Claim 7)
(iii) a Tm value of an oligonucleotide; wherein the higher the Tm value, the higher the rank, (iv) a GC content in a predetermined region of the 5′-end, middle, or 3′-end of an oligonucleotide; wherein the higher the GC content, the higher the rank, (v) the number of consecutive G bases included in an oligonucleotide; wherein the smaller the number of consecutive G bases, the higher the rank,(vi) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when the oligonucleotide forms the homodimer; wherein the smaller the number or proportion, the higher the rank (viii) a length; wherein the shorter the length, the higher the rank. (Claim 9)
wherein the third oligonucleotide candidate group in step (e) is selected by a method comprising the following steps: (e-1) arranging by giving ranks according to at least one of the following arrangement criteria to oligonucleotides having the same start position for designing an oligonucleotide among the oligonucleotides included in the second oligonucleotide candidate group; and (e-2) selecting the highest ranked oligonucleotide from oligonucleotides having the same start position for designing the arranged oligonucleotides, and wherein the arrangement criteria include the following (iii) a Tm value of an oligonucleotide; wherein the higher the Tm value, the higher the rank, (iv) a GC content in a predetermined region of the 5′-end, middle, or 3′-end of an oligonucleotide; wherein the higher the GC content, the higher the rank, (v) the number of consecutive G bases included in an oligonucleotide; wherein the smaller the number of consecutive G bases, the higher the rank,(vi) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when the oligonucleotide forms the homodimer; wherein the smaller the number or proportion, the higher the rank, wherein the larger the free energy value, the higher the rank, and (viii) a length; wherein the shorter the length, the higher the rank. (Claim 10)
(f) providing a primer candidate group for the mutant target nucleic acid sequence by designing primers to amplify a predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence; and (Claim 11)
wherein the primers in step (f) are designed to satisfy at least one of the following conditions: (i) a Tm value of 40-70° C.; (ii) a length of 15-50 bp nucleotides; and (iii) exclusion of a G-run sequence with at least five Gs (Claim 12)
wherein the method further comprises, after step (f), the following steps: (f-1) arranging by giving ranks according to at least one of the following arrangement criteria to primers having the same start position for designing a primer among the primers included in the primer candidate group; and (f-2) selecting the highest ranked primer from primers having the same start position for designing the arranged primers, and wherein the arrangement criteria include the following: (i) the number or proportion of use of a degenerate base and/or universal base introduced into a primer; wherein the smaller the number or proportion of use, the higher the rank (Claim 13)
(iii) the number of (A)n, (T)n, or (C)n mononucleotide run sequences; wherein the smaller the number of the sequences, the higher the rank,(iv) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when a primer forms the homodimer; wherein the smaller the number or proportion, the higher the rank, (vi) a Tm value; wherein the higher the Tm value, the higher the rank, (vii) a GC content; wherein the more the GC content, the higher the rank, and (viii) a length; wherein the shorter the length, the higher the rank (Claim 13)
wherein the position of the nucleotide mutation of interest is expressed as a start position and an end position (Claim 1, Claim 14)
(e) providing a third oligonucleotide candidate group by selecting oligonucleotides from the second oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the third oligonucleotide candidate group is used to detect the nucleotide mutation of interest in the target nucleic acid sequence. (Claim 1, Claim 14)
(vii) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, and (Claim 9)
(vii) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, and (Claim 10)
(v) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, (Claim 13)
(d) providing oligonucleotides satisfying the following selection criteria as a second oligonucleotide candidate group for the mutant target nucleic acid sequence by analyzing matching of the wild-type target nucleic acid sequence with the oligonucleotides included in the first oligonucleotide candidate group for the mutant target nucleic acid sequence wherein the selection criteria include that (i) the number of mismatches between the wild-type target nucleic acid sequence and a predetermined region at the 5′-end, middle or 3′-end of an oligonucleotide is one or more; or (ii) the ratio of mismatches between the wild-type target nucleic acid sequence and an oligonucleotide is a predetermined value or more; and (Claim 1, Claim 14)
(e) providing a third oligonucleotide candidate group by selecting oligonucleotides from the second oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the third oligonucleotide candidate group is used to detect the nucleotide mutation of interest in the target nucleic acid sequence. (Claim 1, Claim 14)
wherein the mismatches (i) in step (d) are minimum mismatches between the wild-type target nucleic acid sequence and the predetermined region of the 5′-end, middle, or 3′-end of the oligonucleotide, and the mismatches (ii) in step (d) are minimum mismatches between the wild-type target nucleic acid sequence and the oligonucleotide. (Claim 8)
wherein the method further comprises, after step (d), d-1) arranging the oligonucleotides included in the second oligonucleotide candidate group by giving ranks according to at least one of the following arrangement criteria:(i) the position of a mismatch in an oligonucleotide as a result of the matching analysis in step (d); wherein the closer the position of the mismatch is to the 5′-end, 3′-end, or middle, the higher the rank, (ii) the number of mismatches of an oligonucleotide as a result of the matching analysis in step (d); wherein the larger the number of the mismatches, the higher the rank, (Claim 9) :
(i) the position of a mismatch in an oligonucleotide as a result of the matching analysis in step (d); wherein the closer the position of the mismatch is to the 5′-end, 3′-end, or middle, the higher the rank, (ii) the number of mismatches in an oligonucleotide as a result of the matching analysis in step (d); wherein the larger the number of the mismatches, the higher the rank, (Claim 10)
(ii) the number of primer patterns generated by the introduction of a degenerate base; wherein the smaller the number of patterns, the higher the rank (Claim 13)
With respect to the limitations of Claims 1, 5, 6, 7, 9, 10, 11, 12, 13, 14, You et al. teaches Sequences can be input in two ways. Sequences in FASTA format can be copied and then pasted to the sequence text box (Figure 1). This approach has a maximum size limit of 256 kb. For a large volume of sequences, a FASTA file can be uploaded to the server and the sequence size limitation only depends on Internet speed and server machine memory. When inputting a FASTA file or a single sequence, a header line starting with ">" is mandatory for each sequence. However, empty lines or spaces within sequences are allowed (Figure 1 and 5). Sequences can be input in two ways. Sequences in FASTA format can be copied and then pasted to the sequence text box (Figure 1). This approach has a maximum size limit of 256 kb. For a large volume of sequences, a FASTA file can be uploaded to the server and the sequence size limitation only depends on Internet speed and server machine memory. When inputting a FASTA file or a single sequence, a header line starting with ">" is mandatory for each sequence. However, empty lines or spaces within sequences are allowed (Figure 1 and 5). As in Primer3Web and Primer3Plus, for any type of primer design, the "{}" pair can be inserted into sequences to specify an included region (for example, excluding the vector sequence fragments on both ends), and the "< >" pair to specify excluded regions. An example is to mask all introns with "< >" to design primers only in exons (Figure 5A). An alternative method to specify excluded regions is to replace the unwanted regions with "N" (Figure 5B) and set the parameter "Max # Ns" as 0. The " []" pair is adopted to specify targets. If multiple targets are set in one sequence, at least one target will be included in the PCR product. It is notable that target masking can be used only for generic primer design in BatchPrimer3. In addition, multiple targets and excluded regions can be specified in a sequence but only one included region is allowed (Program input. It is possible to design two SBE primers, one for each orientation (forward and reverse) For each orientation, all the primer candidates meeting the user-specified primer length range (greater than or equal to the minimum size and less than or equal to the maximum size) are picked. Then the Tm, GC content and quality score of each candidate are calculated. The primer with the highest score is chosen. A pair of SNP flanking primers and SBE primer can be designed in the same module.(SBE primer design) wherein the oligonucleotides in step (c) are designed to have matching or complementary sequences to the predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence. (AS Primer design) The default parameters for PCR primer design were used, and SBE primers were designed in a range of 25 to 35 bases in primer length, 50 to 90°C in Tm and 20 to 80% in GC content. Success rate of SBE primers in PCR amplification was 82.4% (371 out of 450 primer sets), (a) inputting information about a wild-type target nucleic acid sequence and a nucleotide mutation of interest; wherein the wild-type target nucleic acid sequence is a target nucleic acid sequence not comprising the nucleotide mutation of interest, the information includes information about (i) the wild-type target nucleic acid sequence, (ii) a position of the nucleotide mutation of interest occurring in the wild-type target nucleic acid sequence and (iii) wild-type and mutant bases at the position of the nucleotide mutation of interest (Claim 1, Claim 14) and the wild-type and mutant target nucleic acid sequences include forward and reverse wild-type and mutant target nucleic acid sequences, respectively,(Claim 1, Claim 14), (c) providing a first oligonucleotide candidate group for the mutant target nucleic acid sequence by designing oligonucleotides used to detect the nucleotide mutation of interest in a predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence; (Claim 1, Claim 14) wherein the oligonucleotides in step (c) are designed to have matching or complementary sequences to the predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence. (Claim 5) wherein the oligonucleotides in step (c) are designed according to lengths at positions from a predetermined position upstream of the start position of the nucleotide mutation of interest within the mutant target nucleic acid sequence to the end position thereof (Claim 6) wherein the oligonucleotides in step (c) are designed to satisfy at least one of the following conditions: (i) a length of 10-60 nucleotides; (ii) a Tm value of 50-85° C.; (iii) exclusion of a G-run sequence with at least three Gs; and (iv) a GC content of 40% or more in the 5′-end portion (Claim 7) (iii) a Tm value of an oligonucleotide; wherein the higher the Tm value, the higher the rank, (iv) a GC content in a predetermined region of the 5′-end, middle, or 3′-end of an oligonucleotide; wherein the higher the GC content, the higher the rank, (v) the number of consecutive G bases included in an oligonucleotide; wherein the smaller the number of consecutive G bases, the higher the rank,(vi) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when the oligonucleotide forms the homodimer; wherein the smaller the number or proportion, the higher the rank (viii) a length; wherein the shorter the length, the higher the rank. (Claim 9) wherein the third oligonucleotide candidate group in step (e) is selected by a method comprising the following steps: (e-1) arranging by giving ranks according to at least one of the following arrangement criteria to oligonucleotides having the same start position for designing an oligonucleotide among the oligonucleotides included in the second oligonucleotide candidate group; and (e-2) selecting the highest ranked oligonucleotide from oligonucleotides having the same start position for designing the arranged oligonucleotides, and wherein the arrangement criteria include the following (iii) a Tm value of an oligonucleotide; wherein the higher the Tm value, the higher the rank, (iv) a GC content in a predetermined region of the 5′-end, middle, or 3′-end of an oligonucleotide; wherein the higher the GC content, the higher the rank, (v) the number of consecutive G bases included in an oligonucleotide; wherein the smaller the number of consecutive G bases, the higher the rank,(vi) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when the oligonucleotide forms the homodimer; wherein the smaller the number or proportion, the higher the rank, wherein the larger the free energy value, the higher the rank, and (viii) a length; wherein the shorter the length, the higher the rank. (Claim 10) (f) providing a primer candidate group for the mutant target nucleic acid sequence by designing primers to amplify a predetermined region comprising the nucleotide mutation of interest within the mutant target nucleic acid sequence; and (Claim 11) wherein the primers in step (f) are designed to satisfy at least one of the following conditions: (i) a Tm value of 40-70° C.; (ii) a length of 15-50 bp nucleotides; and (iii) exclusion of a G-run sequence with at least five Gs (Claim 12) wherein the method further comprises, after step (f), the following steps: (f-1) arranging by giving ranks according to at least one of the following arrangement criteria to primers having the same start position for designing a primer among the primers included in the primer candidate group; and (f-2) selecting the highest ranked primer from primers having the same start position for designing the arranged primers, and wherein the arrangement criteria include the following: (i) the number or proportion of use of a degenerate base and/or universal base introduced into a primer; wherein the smaller the number or proportion of use, the higher the rank (Claim 13) (iii) the number of (A)n, (T)n, or (C)n mononucleotide run sequences; wherein the smaller the number of the sequences, the higher the rank,(iv) the number or proportion of consecutive nucleotides involved in the formation of a homodimer when a primer forms the homodimer; wherein the smaller the number or proportion, the higher the rank, (vi) a Tm value; wherein the higher the Tm value, the higher the rank, (vii) a GC content; wherein the more the GC content, the higher the rank, and (viii) a length; wherein the shorter the length, the higher the rank (Claim 13)
With respect to the limitations of Claims 1, 9, 10, 13, 14, Untergasser et al. teaches taking a list of region pairs and requires the selected forward and reverse primers to lie in regions specified by at least one pair in the list. Each region is expressed as a starting position, length in bp. three, equally acceptable alternatives for the locations of the forward and reverse primers: Forward primer in the 50 bp region starting at position 100 and reverse primer in the 50 bp region starting at position 300., Forward primer in the 50 bp region starting at position 400 and reverse primer anywhere, Forward primer anywhere and reverse primer in the 30 bp region starting at position 460. (Controlling primer location) ) Primer3 can carry out several kinds of design tasks (discussed later) and also check existing primer pairs for correctness. We focus mainly on discussing Primer3 in the context of designing primer pairs for amplifying a DNA template using PCR. To accomplish this task, Primer3 evaluates the primers and primer pairs according to various constraints and sorts acceptable pairs by a penalty function. It uses ‘branch and bound’ techniques to reduce the search space while still generating the optimal primer pairs according to the penalty function and constraints. (Overview of Primer 3),, wherein the position of the nucleotide mutation of interest is expressed as a start position and an end position (Claim 1, Claim 14) (e) providing a third oligonucleotide candidate group by selecting oligonucleotides from the second oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the third oligonucleotide candidate group is used to detect the nucleotide mutation of interest in the target nucleic acid sequence. (Claim 1, Claim 14) (vii) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, and (Claim 9) (vii) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, and ((Claim 10) (v) a hairpin structure-forming free energy value (ΔG value); wherein the larger the free energy value, the higher the rank, (Claim 13)
With respect to the limitations of Claims 1, 8, 9, 10, 11, 14, Ye et al. teaches the candidate primer pairs are then subject to the specificity checking process. Since Primer3 generates many candidate primer pairs and all of them may need to undergo specificity checking to obtain the specified number of target-specific primer pairs, the entire search process can be very long if each pair is searched with BLAST individually. To solve this problem, we observe that any primer is essentially a sub-region of the PCR template and a single BLAST result using the template as a query should contain alignment information for all primer pairs. As a result, when a user supplies a template sequence to design new primers (the template case), the template itself is submitted for a BLAST search just once, which greatly reduces the total search time. For cases where users submit a pre-existing primer pair to perform specificity checking (the primer-only case), an artificial template sequence is generated for the BLAST search by connecting both primers with a 20 base spacer region of N’s. This ensures that each primer will be treated separately in the BLAST search and thus achieves the equivalent effect of performing a separate BLAST search for each primer. To further minimize the search time, all regions on the template that do not contain candidate primers are masked out to avoid irrelevant BLAST hits. Since all candidate primer locations on the template are established by Primer3 already, amplification targets (amplicons) for all primer pairs can be identified using the single BLAST result above. A primer pair is deemed to be specific only if it has no amplicons on any targets other than the submitted template within the specificity checking threshold specified by the user. Otherwise, it is considered non-specific. In addition to checking for amplicons between the forward and the reverse primers, Primer-BLAST also checks amplicons arising from either primer alone. For example, the forward primer could also act as a reverse primer if it happens to match some regions on the minus strand of the template. The specificity checking module by default uses BLAST search parameters that ensure high sensitivity such that it can detect a target that contains up to 35% mismatches to the primer sequence. The default BLAST expect value cutoff is 30,000 for the primer-only case and it is typically adjusted much higher for the template case (see below). This expect value is 3000 times higher than the standard BLAST program default (the higher the expect value cutoff, the more sensitive the search) and is necessary to ensure detection of targets that have a significant number of mismatches to primers yet are potentially amplifiable in PCR. Other highly sensitive default parameters include a word size of seven (standard BLAST uses 11), 50,000 for the maximum number of database sequences (standard BLAST uses 250) and 1 for match reward to mismatch penalty ratio (standard BLAST uses 1.5). The expect value for a given BLAST match between a primer and a target is roughly proportional to the query sequence length given the same search database , but the query lengths used in the primer-only case and the template case are often very different. Therefore, there can be a large discrepancy in the expect values between the BLAST matches in the two cases, even though the same primer sequences are being aligned. To resolve this issue, we internally adjust the specified expect value cutoff for the template case using the length of the artificial template from the primer-only case as a guide. This ensures that the BLAST results are equivalent between submitting a template and submitting primers only. (Implementation 3-4 paragraphs) Primer-BLAST offers flexible specificity stringency options. Users can specify the number of mismatches that a primer pair must have to unintended targets as well as a 3’ end region where these mismatches must be present. In addition, users can specify the mismatch threshold above which any targets should be ignored (i.e., filtering out targets having too many mismatches to be a concern for non-specific amplification). The default specificity settings are that at least one primer (for a given primer pair) must have two or more mismatches to unintended targets in the last five bases at the 3’ end, and that any targets with six mismatches or more to at least one primer (for a given primer pair) should be ignored. (User interface (d) providing oligonucleotides satisfying the following selection criteria as a second oligonucleotide candidate group for the mutant target nucleic acid sequence by analyzing matching of the wild-type target nucleic acid sequence with the oligonucleotides included in the first oligonucleotide candidate group for the mutant target nucleic acid sequence wherein the selection criteria include that (i) the number of mismatches between the wild-type target nucleic acid sequence and a predetermined region at the 5′-end, middle or 3′-end of an oligonucleotide is one or more; or (ii) the ratio of mismatches between the wild-type target nucleic acid sequence and an oligonucleotide is a predetermined value or more; and (Claim 1, Claim 14) (e) providing a third oligonucleotide candidate group by selecting oligonucleotides from the second oligonucleotide candidate group for the mutant target nucleic acid sequence; wherein the third oligonucleotide candidate group is used to detect the nucleotide mutation of interest in the target nucleic acid sequence. (Claim 1, Claim 14) wherein the mismatches (i) in step (d) are minimum mismatches between the wild-type target nucleic acid sequence and the predetermined region of the 5′-end, middle, or 3′-end of the oligonucleotide, and the mismatches (ii) in step (d) are minimum mismatches between the wild-type target nucleic acid sequence and the oligonucleotide. (Claim 8) wherein the method further comprises, after step (d), d-1) arranging the oligonucleotides included in the second oligonucleotide candidate group by giving ranks according to at least one of the following arrangement criteria:(i) the position of a mismatch in an oligonucleotide as a result of the matching analysis in step (d); wherein the closer the position of the mismatch is to the 5′-end, 3′-end, or middle, the higher the rank, (ii) the number of mismatches of an oligonucleotide as a result of the matching analysis in step (d); wherein the larger the number of the mismatches, the higher the rank, (Claim 9) : (i) the position of a mismatch in an oligonucleotide as a result of the matching analysis in step (d); wherein the closer the position of the mismatch is to the 5′-end, 3′-end, or middle, the higher the rank, (ii) the number of mismatches in an oligonucleotide as a result of the matching analysis in step (d); wherein the larger the number of the mismatches, the higher the rank, (Claim 10)
With respect to the limitations of Claims 1, 13, 14, Rose et al. teaches (vi) The degeneracy is determined at each position of the DNA matrix based on the number of bases found there. As an option, a weight threshold can be specified such that bases that contribute less than a minimum weight are ignored in determining degeneracy. (vii) Possible degenerate core regions are identified by scanning the DNA matrix in the 3′ to 5′ direction. A core region must start on an invariant 3′ nucleotide position, have a length of 11 or 12 positions ending on a codon boundary, and have a maximum degeneracy of 128 (current default). The degeneracy of a region is the product of the number of possible bases in each position. (Materials and Methods, Primer design, 2nd paragraph, A computer-implemented method (Claim 1) A computer readable storage medium containing instructions (Claim 14) (ii) the number of primer patterns generated by the introduction of a degenerate base; wherein the smaller the number of patterns, the higher the rank (Claim 13)
A person havining ordinary skill in the art would easily find Wangkumhang et al. in view of Untergasser et al. in view of You et al. in view of Ye et al. in view of Rose et al. in order to design nucleotides because each work already is in the same field of endeavor of designing nucleotides. A person of ordinary skill in the art would be motivated to see how others had previously designed nucleotides. Primer3 was even designed with the explicit intention of being integrated into other software packages (abstract). Many of the references (such as Wangkumhang et al.) even incorporated Primer3 for the purpose of candidate generation. Applicant just combined prior art to yield a predictable result as each component was already well known in the prior art of nucleotide design. There is a reasonable expectation of success because each part works independently and therefore they are expected to work when combined. Applicant just combined several well known techniques for combining nucleotides in the field. Additionally, person of ordinary skill in the art would recognize how to put together the limited number of combinations primers have been designed in the past in order to optimize that design.
Conclusion
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/C.H.B./Examiner, Art Unit 1687
/Karlheinz R. Skowronek/Supervisory Patent Examiner, Art Unit 1687
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