DETAILED ACTION
Notice of Pre-AIA or AIA Status
1. The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
2. This Application has been reassigned to Examiner Haney in Art Unit 1682.
3. This action is in response to the papers filed May 15, 2026. Any rejections or objections not reiterated herein have been withdrawn. This action is NON-FINAL.
Claims 1, 7, 10-14, 18-26 are currently pending.
Claims 12-14 and 24-26 are withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected invention, there being no allowable generic or linking claim.
Duplicate Claim Warning
4. Applicant is advised that should claim20 be found allowable, claim 21 will be objected to under 37 CFR 1.75 as being a substantial duplicate thereof. When two claims in an application are duplicates or else are so close in content that they both cover the same thing, despite a slight difference in wording, it is proper after allowing one claim to object to the other as being a substantial duplicate of the allowed claim. See MPEP § 608.01(m).
Claim Rejections - 35 USC § 112
5. 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 20-23 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claims 20-21 recite “The method according to claim 10, wherein the solid support in step c) comprises magnetic beads”. This recitation is confusing because step c) of claim 1 does not refer to a solid support. If this recitation was intended to modify the solid support of claim 10 characteristic c) then the claim should be amended to say this.
Claim 22 recites “The method according to claim 10, wherein the temperature in step d) is room temperature. This recitation is confusing because claim 1 only recites steps a), b), and c). If this recitation was intended to modify the temperature of claim 10 characteristic d) then the claim should be amended to say this. Further it is noted that characteristic d) refers to two different temperatures (i) an elevated temperature of 60ºC or above and (ii) a temperature of 40ºC or below. The claim should be amended to clearly indicate that the temperature of 40ºC or below is room temperature.
Claim 23 recites “The method according to claim 10, wherein the sequencing in step i) is next generation sequencing”. This recitation is confusing because claim 1 only recites steps a), b), and c). If this recitation was intended to modify the temperature of claim 10 characteristic d) then the claim should be amended to say this.
Claim Rejections - 35 USC § 103
6. 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.
7. Claims 1, 7, 10, 18, and 20-22 are rejected under 35 U.S.C. 103 as being unpatentable over Conrad (US 2004/0230048 Pub 11/18/2004) in view of Adams (WO 90/01564 Pub 2/22/1990) and Jacobsen (Nucleic Acids Research 2004 Vol 32 No 7 e64).
Conrad teaches a method for the isolation and purification of mRNA, particularly poly(A) RNA (abstract). The method uses isostabilizing salts such as tetramethylammonium chloride (TMAC) and tetraethylammonium chloride TEAC to reduce rRNA carryover during the purification process, thus facilitating the isolation of poly(A) RNA (abstract). Example 2 describes a protocol for poly(A) RNA purification using oligo dT magnetic beads and TMAC. The procedure is as follows:
1. Mix up to 0.5 ml of solution containing 10-1000 µg RNA in water or TE with an equal volume of 4 M TMAC and 1/600th volume of 10% Triton X-100.
2. Remove an aliquot of magnetic beads with a mass equal to that of the total RNA (stocks contain 10 mg magnetic beads per µl of slurry volume; nominal size of 1 µm) and sediment to side of tube using a magnetic stand.
3. Resuspend beads in 2 M TMAC, re-sediment, and repeat.
4. Add the total RNA mixture (in 2M TMAC+0.017% Triton X-10) to the oligo dT magnetic beads that have been equilibrated with 2M TMAC.
5. Incubate the samples at 75ºC for 5 minutes.
6. Incubate the samples with gentle rocking for one hour at room temperature.
7. Sediment the oligo-dT magnetic beads (OdTMB) to the side of the tube using a magnetic stand. Remove supernatant.
8. Resuspend the OdTc in 500 .µl 2 M TMAC. Mix thoroughly. Sediment the OdTMB to the side of the tube using a magnetic stand and discard supernatant.
9. Resuspend the OdTMB in another 500 µl 2 M TMAC. Mix thoroughly. Sediment the OdTMB to the side of the tube using a magnetic stand and discard supernatant.
10. Resuspend the OdTc in 500 µl 0.4 M TMAC. Mix thoroughly. Sediment the OdTMB) to the side of the tube using a magnetic stand and discard supernatant.
11. Repeat step 10.
12. Elute with 2 volumes (100-2001 µl) of the pre-warmed elution buffer (water, TE, or 1 mM Na-citrate). Vortex briefly and pulse spin to collect all of the solution at the bottom of the tube before sedimenting the OdTMB to the side of the tube using a magnetic stand and collecting supernatant.
13. Add 1/10 volume of 3 M sodium acetate to each eluted sample and mix. Add 1 µl (1/200 volume) of glycogen solution to each sample and mix.
14. Add 0.5 ml (2.5 volume) of 100% ethanol to each sample. Mix by inverting several times.
15. Incubate your samples at -20ºC. from 30 minutes to overnight.
16. Centrifuge the samples for 30 minutes at full speed in a tabletop centrifuge. The presence of glycogen will ensure a pellet is visible at the bottom of your tube.
17. Remove the supernatant (we recommend through a drawn-out Pasteur pipette).
18. Add 0.5 ml 70% ethanol to each tube. Vortex briefly to wash the pellets.
19. Centrifuge for 5 min. at full speed.
20. Aspirate the supernatant.
21. Resuspend samples in 1 mM Na-citrate, pH 6.4. The exact volume will depend on the input amount and the analysis planned (see paras 0103-0124).
Conrad teaches that when the samples prepared by examples 1 and 2 were compared by ethidium-staining of a denaturing gel, it could be seen that substantially less rRNA was present than equivalent samples from a standard OdT cellulose procedure, even after two-fold repetitive purifications were performed. Furthermore, on examining the presence of three common mRNA species (βactin, GAPDH, cyclophilin) using the Northern blot technique, the yield was also superior (para 0125). Additionally Conrad teaches that the concentration of the isostabilizing agent in a composition comprising a sample and a poly(T) nucleic acid is between about 1.0M and about 3.0M (para 0015).
Thus Regarding Claim 1 Conrad teaches a method for isolating a poly(A) nucleic acid having a single stranded poly(A) nucleic acid stretch of consecutive adenine base residues from a nucleic acid containing sample comprising poly(A) nucleic acids having a single stranded poly(A) nucleic acid region, comprising:
(a) contacting the sample with
i. a quaternary ammonium salt (TMAC or TEAC), and
ii. a capture probe capable of hybridizing to the poly(A) stretch of the poly(A) nucleic acids (the OdTMBs) (see step 4);
(b) incubating after the contacting to form nucleic acid-hybrids between the poly(A) nucleic acids and the capture probe (see steps 5 and 6); and
(c) separating the formed hybrids from the remaining sample (see steps 7-10).
Conrad teaches that the quaternary ammonium salt is at a concentration of 0.25 M to 1.25 M after the contacting.
Regarding Claim 7 Conrad teaches a method wherein the poly(A) nucleic acid is poly(A) RNA (Example 2).
Regarding Claims 10, 20, and 21 Conrad teaches a method wherein the capture probe is bound to a solid support wherein the solid support is a magnetic bead (see step 4).
Regarding Claims 10 and 22 Conrad teaches a method wherein the incubating is at elevated temperature of 60°C or above and is subsequently incubated at a temperature of 40°C or below to allow hybridization of the poly(A) RNA to the capture probe, wherein the temperature of 40ºC or below is room temperature (see steps 5 and 6).
Regarding Claim 18 Conrad teaches a method wherein the poly(A) RNA is total RNA (see step 4).
Conrad does not teach a method wherein the sample is contacted with a sodium salt AND a quaternary ammonium salt as a single solution or separately in any order with the sample. Conrad does not teach a method wherein the sodium salt is NaCl and the NaCl is in a concentration of 75 mM to 250 M before the contacting or at a concentration of 37.5 mM to 125 mM after the contacting (clm 1). Finally Conrad does not teach a method wherein the quaternary ammonium salt is TMAB, TMAN, or TEAB (clm 1).
However Adams teaches a method for assaying the presence of target nucleic acids, the method comprising contacting a hybridization medium containing target nucleic acids with a dipstick comprising a solid surface having a multiplicity of discrete regions with different nucleic acid probes covalently bound thereto (page 5, lines 9-14). Adams discloses a common hybridization solution that comprises sodium chloride (page 17, lines 17-21). Adams also teaches that to control the effect of different GC contents between the nucleic acid reactants, the methods may take place in a hybridization buffer comprising an ammonium salt selected from a trialkylammonium salt and a tetraalkylammonium salt. Adams teaches that preferred tetraalkylammonium salts are tetramethylammonium chloride and tetraethylammonium chloride. Adams teaches that the anionic portion of the salt may be chloride or bromide. The preferred concentration of the ammonium salt is between 2 and 3.5 moles per liter (page 6 line 26 to page 7 line 3).
Additionally Jacobsen teaches a method for the direct isolation of poly(A) RNA using locked nucleic acid-oligo(T) capture. Jacobsen teaches that the LNA 2.T affinity probe was highly efficient in isolation of poly(A) RNA in a low salt concentration range of 50-100 mM NaCl in poly(A) binding buffer (abstract).
Accordingly, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Conrad by using TMAB or TEAB as the quaternary ammonium salt as suggested by Adams. In the instant case Conrad teaches that TMAC and TEAC negate the difference in the hydrogen bonding between A-T/U and G-C base pairs (para 0014). Adams teaches that tetraalkylammonium salts control the effect of different GC contents between the nucleic acid reactants. Adams teaches that preferred tetraalkylammonium salts are tetramethylammonium chloride and tetraethylammonium chloride. Adams teaches that the anionic portion of the salt may be chloride or bromide (page 6). The claim would have been obvious because the substitution of one quaternary ammonium salt (TMAC or TEAC of Conrad) for another (TMAB or TEAB of Adams) would have yielded predictable results to one of ordinary skill in the art at the time of the invention. Further it would have been obvious to modify the method of Conrad by contacting the sample with NaCl AND TMAB or TEAB as suggested by Adams and Jacobsen. Adams teaches hybridization solutions which comprise NaCl and that TMAB or TEAB may be added to hybridization solutions to control the effect of different GC contents. Jacobsen teaches that the LNA 2.T affinity probe was highly efficient in isolation of poly(A) RNA in a low salt concentration range of 50-100 mM NaCl in poly(A) binding buffer (abstract). One of skill in the art would have been motivated to contact a sample with NaCl AND TMAB or TEAB because the prior art teaches highly efficient capture of poly(A) RNA in NaCl solutions and control of the effect of different GC contents by using TMAB or TEAB.
Further, to have determined the optimum concentration of NaCl and TMAB or TEAB would have been obvious to one of ordinary skill in the art and well within the skill of the art. As discussed in MPEP 2144.05(b), “(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, 105 USPQ 233, 235 (CCPA 1955).
MPEP 2144.05(b):
“Generally, differences in concentration or temperature will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration or temperature is critical. "[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)”
“A particular parameter must first be recognized as a result-effective variable, i.e., a variable which achieves a recognized result, before the determination of the optimum or workable ranges of said variable might be characterized as routine experimentation. In re Antonie, 559 F.2d 618, 195 USPQ 6 (CCPA 1977).”
8. Claims 10, 11, 19, and 23 are rejected under 35 U.S.C. 103 as being unpatentable over Conrad (US 2004/0230048 Pub 11/18/2004) in view of Adams (WO 90/01564 Pub 2/22/1990) and Jacobsen (Nucleic Acids Research 2004 Vol 32 No 7 e64) as applied to claim 1 above and in further view of Ozsolak (Nature Reviews Genetics Vol 12 2/2011 pages 87-98).
The teachings of Conrad, Adams, and Jacobsen are presented above.
The combined references do not teach a method further comprising sequencing the poly(A) RNA (clms 10, 11, 19). The combined reference do not teach a method wherein the sequencing is next generation sequencing (clm 23).
However Ozsolak teaches a method of sequencing poly(A) RNA using a next generation sequencing method (see Fig 4, page 94-95).
Accordingly, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Conrad, Adams, and Jacobsen by further sequencing the poly(A) RNA using a next generation method as suggested by Ozsolak for the benefit of being able to accurately measure gene expression. One of skill in the art would have been motivated to use a next generation sequencing method because it is high throughput and high speed.
6. Any inquiry concerning this communication or earlier communications from the examiner should be directed to AMANDA HANEY whose telephone number is (571)272-8668. The examiner can normally be reached Monday-Friday, 8:15am-4:45pm EST.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Wu-Cheng Shen can be reached at 571-272-3157. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/AMANDA HANEY/Primary Examiner, Art Unit 1682
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