DETAILED ACTION
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
The amendment filed December 22, 2025, has been received and entered.
Claims 8 and 10 are cancelled.
Claims 1-7, 9, 11, and 12 are pending and examined on the merits.
Notice Re: Prior Art Available Under Both Pre-AIA and AIA
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.
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.
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 1-7, 9, 11, and 12 are rejected under 35 U.S.C. 103 as being unpatentable over Ma (CN 103525866. Listed on IDS filed 12/5/24. Translation cited below, annexed to the Extended European Search Report of EP 21867584 of patent family of instant application) in view of Domanico (US 2009/0246859. Listed on IDS filed 12/5/24), Engebrecht (Current Protocols in Immunology. 1998. Chapter 10: Unit 10.3. pages 10.3.1-10.3.10. Previously cited) and Ciccolini (Biotechnology and Bioengineering. 1998. 60(6): 768-770. Previously cited)
Ma discloses a method for preparing a plasmid (paragraph [0014] of Translation). The method comprises lysing a suspension of Escherichia coli to obtain an E. coli lysate, wherein the E. coli contains a plasmid (paragraphs [0015] and [0017] of Translation). The lysing of the E. coli suspension comprises mixing the E. coli suspension with a lysis solution containing 0.2 M NaOH and 1% SDS at 15-30 rpm (paragraphs [0017] and [0033] of Translation). This meets limitations of step (a) of instant claim 1 since it is directed to adding a lysis solution to a bacterial cell suspension to obtain a first mixture. The treated bacterial cell suspension is directed to the ‘preliminary bacterial cell lysate’ of instant claim 1. Further still, the lysis solution containing 0.2 M NaOH and 1% SDS of Ma is an ‘alkaline lysis buffer’ as instantly claimed because this same solution is taught in Engebrecht, specifically a 0.2 M NaOH solution containing 1% (w/v) SDS, for alkaline lysis (page 768, right column, first paragraph; page 769, left column, third paragraph of Engebrecht).
Then, the E. coli suspension treated by the lysis solution is mixed with a potassium acetate solution and stirred at 15-30 rpm in order to obtain the E. coli lysate (paragraphs [0017] and [0034] of Translation). Potassium acetate is directed to an acetate buffer, meeting the buffer limitations of instant claims 1 and 11. Therefore, this step of the lysing taught in Ma meets limitations of step (b) of instant claim 1 since it is directed to adding an acetate buffer (the potassium acetate solution) to the preliminary bacterial cell lysate to obtain a second mixture.
The mixing steps of Ma differ from the claimed invention in that they are accomplished by stirring, as opposed to shaking using a first shaker (for mixing with the lysis solution) and a second shaker (for mixing with the potassium acetate solution) as claimed.
However, Domanico discloses methods for gently lysing and solubilizing cells (abstract), as well as methods for purifying plasmid DNA from host bacterial cells (paragraph [0041]). In one example for isolating plasmid DNA, a lysis solution was added to bacterial host cells having a plasmid, and then shaken to thoroughly mix the cells in the lysis solution (paragraph [0120]). The data of the example indicated that mixing the cells with lysis solution can be accomplished by shaking plates on a plate shaker (paragraph [0121]).
Before the effective filing date of the claimed invention, it would have been obvious to the person of ordinary skill in the art to perform shaking using a shaker, instead of stirring, under the same rotational speed of 15-30 rpm, for each of the treatments with the lysis solution and the potassium acetate solution, such that a first shaker is used for shaking with the lysis solution and a second shaker is used for shaking with the potassium acetate solution, when performing the method of Ma for the predictable result of mixing the lysis solution with the E. coli suspension and mixing the potassium acetate solution with the E. coli suspension that had been treated with the lysis solution. It would have been a matter of simple substitution of one technique for another of mixing solutions with a cell suspension for the purpose of preparing a plasmid from bacterial cells. It would have been obvious to use the same rotational speed for shaking as used in stirring since it was expressly taught in Ma for their invention to effect mixing. Therefore, Ma in view of Domanico renders obvious shaking the ‘first mixture’ (resulting from adding the lysis solution to the E. coli cell suspension) using a first shaker at a rotation speed of 15-30 rpm which overlaps with the range of ‘from about 10 rpm to about 30 rpm’ of step (a) of instant claim 1, and shaking the ‘second mixture’ (resulting from adding the potassium acetate solution to the E. coli suspension treated with the lysis solution) using a second shaker at a rotation speed of 15-30 rpm which overlaps the range of ‘from about 10 rpm to about 70 rpm’ of step (b) of instant claim 1.
Ma in view of Domanico differs from the claimed invention in that they do not expressly disclose that each of the shaking steps at 15-30 rpm is for about 1 minute to about 10 minutes, followed by incubation after each (that is, incubation after the shaking of the E. coli suspension and the lysis solution, and incubation after the shaking of that suspension and the potassium acetate solution).
Engebrecht is a review of methods for preparing bacterial plasmid DNA free from chromosomal DNA (page 10.3.1, third paragraph). One method is a miniprep procedure in which plasmid-containing bacteria are lysed by treatment with a solution containing SDS, which denatures bacterial proteins, and NaOH, which denatures chromosomal and plasmid DNA (page 10.3.1, last paragraph). The mixture is neutralized with potassium acetate, causing the covalently closed plasmid DNA to reanneal rapidly, as well as causing the precipitation of most of the chromosomal DNA and bacterial proteins (page 10.3.1, last paragraph). The steps of the procedure are described on page 10.3.2, with the materials set forth in the paragraph bridging pages 10.3.1 and 10.3.2. In particular, an NaOH/SDS solution comprising 0.2 N NaOH and 1% SDS is added to a suspension of plasmid-bearing E. coli cells, mixed by tapping the tube with a finger, and placed on ice for 5 minutes. Then, a potassium acetate solution is added and vortexed at maximum speed for 2 seconds to mix. The mixture is then placed on ice for 5 minutes.
Ciccolini discloses the laboratory preparation of plasmid DNA from Escherichia coli carrying the plasmid vector (page 768, paragraph bridging left and right columns). It includes an alkaline lysis reaction which normally starts by addition and gentle shaking of a known volume of cell suspension with a solution of sodium hydroxide (NaOH) containing sodium dodecyl sulfate (SDS) (page 768, right column, first paragraph). The alkaline rich environment of the mixture causes the denaturation of low molecular weight plasmid DNA and high molecular weight chromosomal DNA (page 768, right column, first paragraph). Following a period of about 5 minutes, which is thought to be sufficient for the lysis reaction and denaturation to be complete, the mixture is neutralized by the addition of a concentrated, chilled solution of potassium acetate (page 768, right column, first paragraph).
The study of Ciccolini investigated the time course of the SDS-alkaline lysis of recombinant E. coli cell suspensions (abstract). The course of reaction was followed by carrying out the lysis operation in the cup of a coaxial cylinder rheometer (page 769, left column, second paragraph). In particular, the lysis reaction experiment was started by addition of 0.2 M NaOH containing 1% (w/V) SDS, and recording the viscosity continuously as a function of time for up to 10-12 minutes (page 769, paragraph bridging left and right columns). The increase in viscosity reached a peak value between 80 and 120 seconds, i.e. between about 1.3 and 2 minutes (paragraph bridging pages 769 and 770). Ciccolini states that the time required for viscosity to rise to its final peak value evidently corresponded to the time for the completion of chromosomal DNA denaturation and maximal entanglement (80-120 s), and the gradual decrease in viscosity following the peak reflected the expected shear degradation of the denatured chromosomal DNA (page 770, right column, third paragraph). Also, the apparent viscosity falls to a steady value after about 200-400 seconds, i.e. about 3.3-6.7 minutes (page 770, left column, first paragraph).
Before the effective filing date of the claimed invention, it would have been an obvious matter of routine optimization to vary the length of time of the shaking at 15-30 rpm for each treatment with the lysis solution and the potassium acetate solution using the respective first and second shakers when performing the method rendered obvious by Ma in view of Domanico, specifically varying the shaking time to about 1 minutes to about 10 minutes, or about 3 minutes to about 8 minutes for the two treatments; also, varying the shaking time to about 1 minute to about 8 minutes for the treatment with the lysis solution. This would have been obvious because the skilled artisan would have recognized that the length of time of the shaking would have affected the extent of the denaturation of the chromosomal and plasmid DNA by the lysis solution, the neutralization by the potassium acetate for reannealing the covalently closed plasmid DNA, and the precipitation of the chromosomal DNA and bacteria, based on the teachings of Engebrecht and Ciccolini, including the teaching of a reaction time of 1.3-3 minutes in Ciccolini which would have served as a starting point for optimization of the shaking time for the treatment with the lysis solution. It is noted that “[W]here the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.” In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955).
Further still, before the effective filing date of the claimed invention, it would have been obvious to the person of ordinary skill in the art to incubate the mixtures after each treatment with the lysis solution and the potassium acetate solution when performing the method rendered obvious by Ma in view of Domanico, Engebrecht, and Ciccolini. One of ordinary skill in skill in the art would have been motivated to do this because such incubations after mixing were well known, as indicated in Engebrecht and Ciccolini, for a method of preparing plasmids from plasmid-containing E. coli using the same treatment with a lysis solution comprising 0.2 M NaOH and 1% (w/v) SDS followed by potassium acetate solution. There would have been a reasonable expectation of obtaining an E. coli lysate for plasmid preparation, as sought by Ma, by this modification because these incubation periods were included in treatment with the same chemicals for plasmid release and recovery in Engebrecht and Ciccolini.
As such, Ma in view of Domanico, Engebrecht, and Ciccolini renders obvious instant claims 1, 3, 4, 7, and 11.
Regarding instant claim 2, the shaking at a rotation speed of 15-30 rpm for the mixing with the lysis solution, as rendered obvious by the references, overlaps the range of instant claim 2. Thus, instant claim 2 is rendered obvious.
Regarding instant claim 5, the references render obvious shaking at a rotation speed of 15-30 rpm for the mixing with the potassium acetate solution. Rotation speeds in the higher end of this range (e.g., 29 rpm, 30 rpm) fall within the range of instant claim 5 (29 rpm and 30 rpm each are directed to ‘about 30 rpm’). Thus, instant claim 5 is rendered obvious.
Regarding instant claim 6, the references differ from the claimed invention in that they do not expressly disclose that the shaking using a second shaker of the E. coli suspension (which had been treated with the lysis solution) with the potassium acetate solution is at a rotation speed ranging from about 40 rpm to about 50 rpm. Engebrecht teaches mixing with the NaOH/SDS solution by tapping tube finger, whereas the mixing with the potassium acetate solution is by vortex at maximum speed (page 10.3.2). Before the effective filing date of the claimed invention, it would have been a matter of routine optimization to use a higher rotation speed of the shaking using a second shaker of the potassium acetate solution with the E. coli suspension (that had been treated with the lysis solution), including optimization to a rotation speed ranging from about 40 rpm to about 50 rpm, when performing the method rendered obvious by Ma in view of Domanico, Engebrecht, and Ciccolini, because the skilled artisan would have recognized that the rotation speed would have affected the mixing which is turn affects the extent of neutralization by the potassium acetate solution to cause the covalently closed plasmid DNA to reanneal and precipitation of the chromosomal DNA and bacterial proteins (see page 10.3.1, last paragraph of Engebrecht). Moreover, it would have been obvious to use a higher rotation speed of the shaking using a second shaker of the potassium acetate solution with the E. coli suspension because increased agitation was taught in Engebrecht for the treatment with potassium acetate solution as compared with the treatment with the lysis solution (solution containing SDA and NaOH). It is noted that “[W]here the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.” In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955). Therefore, instant claim 6 is rendered obvious.
Regarding instant claim 9, as pointed out above, the lysis solution of Ma contains 0.2 M NaOH and 1% SDS (paragraphs [0017] and [0033] of Translation). This same solution is taught in Engebrecht, specifically a 0.2 M NaOH solution containing 1% (w/v) SDS, for alkaline lysis (page 768, right column, first paragraph; page 769, left column, third paragraph). Therefore, the lysis solution of Ma is directed to an alkaline lysis buffer comprising 1% (w/v) SDS and 200 mM NaOH. As such, instant claim 9 is rendered obvious.
Regarding instant claim 12, as pointed out above, the method of Ma is a method of preparing a plasmid. After obtaining the E. coli lysate through treatments with the lysis solution and the potassium acetate solution (paragraphs [0015] and [0017] of Translation), Ma teaches steps of microfiltration of the E. coli lysate to purify the E. coli lysate, subjecting the E. coli lysate to ultrafiltration, and isolating the plasmid from the ultrafiltrated lysate (paragraph [0015] of Translation). Thus, instant claim 12 is rendered obvious.
Response to Arguments
Applicant’s arguments, filed December 22, 2025, with respect to the objection to claim 9 have been fully considered and are persuasive. In particular, the objection has been overcome by the amendment to claim 9. Therefore, the claim objection has been withdrawn.
However, Applicant’s arguments are unpersuasive with respect to the rejection under 35 U.S.C. 103 which has been modified as necessitated by the claim amendments. Applicant argues that the speeds in RPM for the stirrers (Ma) and shakers (in the claimed method) are different, based on discussion indicating that in magnetic stirrers, for example, the rotational speed is of the magnetic bar in units of RPM which indicates how many times the magnetic bar completes a full turn in one minute, whereas the RPM for a 3D shaker, for example, describes the number of complete motion cycles the shaker platform completes in one minute. However, as set forth in the rejection, it would have been a matter of simple substitution of one technique for another of mixing solutions with a cell suspension for the purpose of preparing a plasmid from bacterial cells. Though the RPM disclosed in Ma is for shaking instead of stirring, the skilled artisan would have recognized that the RPM disclosed in Ma is directed to the RPM of the motion of the cell suspension that applies to the substitution with a shaker in order to effect mixing. That is, the speeds in RPM for the stirrers in Ma do speak to the motion of the cell suspension when using a shaker, specifically the RPM of the shaker.
Additionally, Applicant points out that the lysis solution of Domanico as described in its Table 3 is not an alkaline lysis buffer taught by Ma, and that, instead, Domanico teaches that alkaline lysis does not provide quality plasmid DNA, citing paragraphs [0005] and [0060] of Domanico. Therefore, Applicant asserts that a skilled artisan would not have been motivated to combine Ma and Domanico to arrive at the claimed methods where an alkaline lysis solution is used in combination with shaking at specific speed ranges for a specific time period. However, Domanico is cited for demonstrating that bacterial cells can be mixed with a lysis solution by shaking plates on a plate shaker (paragraph [0121]). Though Ma and Domanico use different lysis solutions, the use of a shaker as taught in Domanico is applicable to Ma as a technique for mixing a solution with a cell suspension for the purpose of preparing a plasmid from bacterial cells. A shaker is suitable for the purpose of mixing, regardless of the contents of the materials being mixed.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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Sef
/SUSAN E. FERNANDEZ/Examiner, Art Unit 1651
/DAVID W BERKE-SCHLESSEL/Primary Examiner, Art Unit 1651