SOURCES SOUGHT
A -- Generation of a Conditional Knockout Mouse Model with Reporter
- Notice Date
- 8/4/2014
- Notice Type
- Sources Sought
- NAICS
- 541711
— Research and Development in Biotechnology
- Contracting Office
- Department of Health and Human Services, National Institutes of Health, National Cancer Institute, Office of Acquisitions-Treatment and Support Branch, Bldg 244, Room 112, Frederick, Maryland, 21702
- ZIP Code
- 21702
- Solicitation Number
- HHSN-NIH-NCI-SBSS-TSB-44440-11
- Archive Date
- 8/26/2014
- Point of Contact
- Brenda Oberholzer, Phone: 301-624-8750
- E-Mail Address
-
oberholzerb@mail.nih.gov
(oberholzerb@mail.nih.gov)
- Small Business Set-Aside
- N/A
- Description
- Generation of a Conditional Knockout Mouse Model with Reporter This is a Small Business Sources Sought notice. This is NOT a solicitation for proposals, proposal abstracts, or quotations. The purpose of this notice is to obtain information regarding: (1) the availability and capability of qualified small business sources; (2) whether they are small businesses; HUBZone small businesses; service-disabled, veteran-owned small businesses; 8(a) small businesses; veteran-owned small businesses; woman-owned small businesses; or small disadvantaged businesses; and (3) their size classification relative to the North American Industry Classification System (NAICS) code for the proposed acquisition. Your responses to the information requested will assist the Government in determining the appropriate acquisition method, including whether a set-aside is possible. An organization that is not considered a small business under the applicable NAICS code should not submit a response to this notice. This requirement is assigned a code of 541711 in the North American Industry Classification System (NAICS), and the size standard for such requirements is 500 Employees. The National Center for Complementary and Alternative Medicine (NCCAM), is anticipating to award a Firm-Fixed Price purchase order to generate a knockout-first Fgf13 mouse model expressing a fluorescent reporter, with the option to rescue the conditional Fgf13 allele. The targeting strategy will specifically account for the exon and intron structure of the Fgf13 gene, with an emphasis on supporting knockout-first with fluorescent reporter expression, as well as the wild-type expression of the floxed allele and the final production of a complete null allele following Cre-mediated recombination. To accomplish this goal, a special FRT flanked trap cassette containing a splice acceptor, 2A peptide, a fluorophore, polyA and Neomycin selection cassette followed by a LoxP site will be inserted into the intron upstream of the target exon(s). A single LoxP site will be inserted downstream of the target exon(s). The mice generated in this project will be used to perform electrophysiological, imaging, and behavioral experiments in my lab, that will aim at elucidating the functional roles of Fgf13 in the modulation of pain. Services needed and expected deliverable schedules: Services Deliverables Expected Deliverable Schedules 1. Cloning and construction of a targeting vector to generate an Fgf13 knockout with YFP expression first and a rescue to conditional allele of Fgf13 • Complete bioinformatics analysis of Fgf13 gene structure and sequence content. • Technical review of any previous targeting history of the Fgf13 gene. • Presentation of all relevant targeting strategies that meet our experimental objectives. • Final Targeting Strategy approved by COR. • Screening strategy design for PCR and Southern blot analysis. • Confirmation of genomic BAC clone containing the Fgf13 gene. • Cloning and confirmation of homology arms. • Insertion and confirmation of cassette trap containing a splice acceptor, 2A peptide, YFP, polyA and Neomycin selection cassette flanked by FRT sites, as well as LoxP site insertions around target region.. • Sequencing of targeting vector. • Linearization of DNA for electroporation. Deliverabl: Targeting Vector Expected delivery: ~2-3 months 2. Electroporation and tissue culture of embryonic stem (ES) cell clones • Culture and expansion of ES cells • Electroporation of ES cells with targeting vector • Positive/negative selection of ES cells • Isolation of 200-300 ES cells per electroporation • Duplication and freezing of ES cells • Isolation of DNA for screening of ES cells Estimated Delivery: ~1.5 months 3. Screening of ES cell clones to identify the positive clones • PCR and Southern blotting for identification and confirmation of positive clones. • Assessment of the genomic stability of ES cell clones by determination of percentage of euploid cells to ensure that ES cells of high genomic stability are used for injection and mouse production. • Expansion of positive ES cell clones and preparation for microinjection. Deliverable: Positive ES Clone Estimated Delivery: ~1 month 4. Microinjection of blastocysts with positive clones and birth of chimeras • Preparation of embryos and foster mice • Microinjection of targeted ES clones into blastocysts or laser assisted injection of 8-cell stage embryos • Implantation of blastocysts/8-cell embryos. Deliverable: Chimera Estimated Delivery: ~1 month 4. Animal husbandry to generate F1 mice • Maintenance and care of recipient females and subsequent weaning of chimeras • Breeding of male chimeras with C57Bl/6N wild type females • Weaning of the F1 generation. • F1 genotyping via tail biopsy and DNA extraction. • PCR analysis and confirmation of project specific modifications. Chimeras and Deliverable: F1 mice Estimated Delivery: ~3-4 months 5. Delivery of mice • Guarantee the delivery of a minimum of 2 live heterozygous mice at no additional cost. The mice delivered will be Fgf13 knockout mice with fluorescent reporter expression. These mice can then be used to rescue the conditional allele by mating to FLP to delete the trap cassette and fluorophore. • An approved courier for shipment of live mice, that ensures timely and climate controlled and sensitively handled delivery must be used. Additional Specific Service Needs: • Guarantee the generation of the full service mouse models. Trouble shooting at no additional cost if needed. • Flexibility that will allow us to design our own targeting vector or to choose from the proprietary vectors of the company. • Project information must be kept confidential. All intellectual property belongs to NCCAM. The contractor cannot re-make or re-sell this specific mouse model. • Delivery of end product (F1 mice) within a year. • Monthly comprehensive updates. • Post project consultations to ensure client is able to breed and genotype mice. Delivery Information: Yarimar Carrasquillo National Institutes of Health Building 35A, Room 1E410 35 Convent Drive Bethesda, MD 20814 To be deemed capable of providing the current need the offeror must submit a written capability statement. Organizations that submit capability statements in response to this notice will be evaluated against the following technical areas of experience and expertise: A. Organizational Experience and Capability Demonstrate the organizational experience and capability as required for the performance of the tasks stated in the SOW. Specifically, addressing the following two areas: (1) Demonstrate pertinent background experience and qualifications of the organization particularly with projects involving design, cloning and construction of targeting vectors similar to the one described in the SOW as well as the successful completion of the generation of knockout mice with fluorescent reporters and mice carrying conditional floxed alleles (i.e. How long has the company been in business; how many mouse models have been completed to date; what percentage of attempted mouse models has the company been able to successfully complete; what percentage of mouse models are fully completed within a year). (2) A special notation should be made of similar or related programs performed for the Government including documentation with reference to applicable contract numbers and supervising agencies. B. Personnel Personnel described must have experience with the design, cloning and construction of targeting vectors used to generate knockout-first with fluorescent reporter, then a floxed allele to generate conditional knockouts. In addition, the team shall have PhD level molecular biologists, cytogeneticists and microinjectionists with at least 3 years of experience in the generation of knockout mice, as well as animal husbandry staff all specialized to their specific part in generating mouse models. C. Technical Approach A detailed Technical Approach that demonstrates a clear understanding of the SOW with discussions of a) cloning and construction of targeting vectors; b) electroporation and tissue culture of ES cell clones; c) screening of ES cell clones; d) microinjection of blastocysts; e) animal husbandry to generate F1 mice and e) packaging, labeling and shipping method of finished products. D. Facilities and Equipment Demonstrate the availability of resources, equipment, and facilities as required for the performance of the tasks stated in the SOW. Specifically, addressing the following two areas: (1) Hybrid (129/C57BL/6) ES cells lines shall be available. (2) Animal Assurance through the Office of Laboratory Animal Welfare (OLAW). An IACUC approval will be required prior to commencing work with animals. If any responsible source believes it can perform the requirements they may submit their capability statements via email (PDF format; 15 Page Limit, these pages exclude resumes) for consideration by NCI to Brenda Oberholzer, Contracting Officer, at the e-mail addresses oberholzerb@mail.nih.gov by 12:00 PM ET, August 11, 2014. No collect calls or facsimile transmissions will be accepted. Respondents must include DUNS number, organization name, address, point of contact, and size and type of business (e.g., 8(a), HUBZONE, etc) pursuant to the applicable NAICS code. Disclaimer and Important Notes: This notice does not obligate the Government to award a contract or otherwise pay for the information provided in response. The Government reserves the right to use information provided by respondents for any purpose deemed necessary and legally appropriate. Any organization responding to this notice should ensure that its response is complete and sufficiently detailed to allow the Government to determine the organization's qualifications to perform the work. Respondents are advised that the Government is under no obligation to acknowledge receipt of the information received or provide feedback to respondents with respect to any information submitted. After a review of the responses received, a pre-solicitation synopsis and solicitation may be published in Federal Business Opportunities. However, responses to this notice will not be considered adequate responses to a solicitation(s). Confidentiality: No proprietary, classified, confidential, or sensitive information should be included in your response. The Government reserves the right to use any non-proprietary technical information in any resultant solicitation.
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