About The DNA Synthesis Call
The mission of the DNA Synthesis Science Program is to enable users to perform state-of-the-art functional genomics research and to help them translate genomic information into biological function. Our strategies are to provide users:
- Access to large-scale DNA synthesis and assembly capabilities
- Access to integrated genome technologies and informatics pipelines — from sequencing data generation and database mining including JGI genome databases (Genome portal; bacteria: IMG; fungi: MycoCosm; plants: Phytozome), to construct design, DNA synthesis, microbial strain engineering, transcriptomics, and high-throughput functional characterization.
Additionally, our value proposition is to enable users to study the function of genes that are otherwise difficult to source from environmental samples, including uncultured microbes and higher plants. For consortium-type projects, we can help coordinate the consortiums to publish articles on the highest impact journals.
Proposals to access these capabilities may be submitted to the JGI at any time in the form of brief white papers. Proposals are reviewed twice a year for scientific merit, DOE relevance, proposed usage by the applicant/community, and the applicant’s track record. Applicants are encouraged to review Synthetic Biology Internal Review Process guidelines, as insufficient information will delay or potentially defer approval for the proposal.
For questions about whether your project is appropriate, or for program specifics or technical guidance, please contact Yasuo Yoshikuni (program head) and Miranda Harmon-Smith (project manager). For questions about the application process, please contact Miranda Harmon-Smith.
The current call for proposals accepts applications in three areas, as described below:
1. Synthesis of genes and pathways for functional characterization. DNA synthesis technology has advanced to the point that now it allows access to many genes and pathways within diverse, large, and rapidly expanding sequence spaces for heterologous expression. The technology could facilitate discovery and characterization of gene products with novel and/or desired biochemical function at an unprecedented scale. To support this work, the JGI solicits proposals requesting large-scale DNA synthesis and construct assembly of genes and pathways for functional characterization. Synthesis and assembly are carried out in the DNA Synthesis Platform, which offers the synthetic biology product sizes shown below, with the estimated cycle times listed.
- Short size constructs (<1 kb): ~80 calendar days
- Medium size constructs (1-5 kb): ~110 calendar days
- Large size constructs (5-10 kb): ~ 170 calendar days
- Very large size constructs (10-50 kb): ~210 calendar days
The JGI also supports sequence data mining through the JGI’s genome portals and selection of target genes and pathways for synthesis. These selections are made with criteria including phylogenetic distributions and Pfam compositions in pathways. All constructs are synthesized and assembled into user-defined plasmids, sequence validated, and transformed into an E. coli strain before shipment to users. The products are delivered to users as glycerol stocks.
The program accepts proposals from single investigators and consortiums. A consortium must include participation of principal investigators from at least three different institutions and a plan to disseminate materials synthesized by the program to the scientific community. A single investigator can request a total of 50 to 500 kbp of DNA synthesis capacity per proposal. A consortium can request large-scale synthesis capacity (50 to 1,500 kbp in total per proposal).
2. Synthesis of combinatorial pathway libraries for fast-track metabolic engineering. Metabolic engineering traditionally approaches strain development stepwise, requiring cycles of construct design, building, and characterization. However, pathway efficiency may be optimized more efficiently if multiple pathway variations are tested simultaneously. The JGI recently developed the ability to efficiently build more than 400 pathway variations comprising promoters, RBSs, gene variations, and terminators with different strengths and/or activities. Each proposal may request up to 4,000 kbp of DNA synthesis capacity. The JGI will also help identify a panel of each pathway component and design final constructs. All constructs are assembled using type II restriction-enzyme-based technologies (e.g., golden gate assembly) into user-defined plasmids and are transformed into E. coli strains before shipment to users; no sequencing validations will be performed for the constructs. The products are delivered to users as glycerol stocks.
3. Synthesis of sgRNA libraries. CRISPR/CAS9 technologies facilitate genome editing (deletion, insertion, and point mutation) and transcriptional regulation (activation and inhibition) of any genes of interest at an unprecedented scale and are therefore useful for high throughput functional genomics studies. Each proposal may request up to four libraries comprising up to 12,000 sgRNA sequences. The JGI can help design sgRNA sequences based on the genome sequences of targeted microbes. All sgRNA constructs are synthesized, cloned into user-defined plasmids, and transformed into an E. coli strain as pools. The quality of these libraries is evaluated with sequencing-based analysis using MiSeq before shipment to users. The JGI will create at least 10 replicates of each library to minimize variations in experiments, and will deliver them to users as glycerol stocks. The sgRNA libraries are purified and subsequently transformed into the targeted microbes. These transformants are subjected to user-defined functional screenings. The JGI can further evaluate enriched sgRNA libraries with sequencing-based analysis using MiSeq.
DNA Synthesis Science proposals are accepted on a continuous basis and will be reviewed twice a year. Submission deadline for reach review process is listed below. Letters of intent are not required.
- July 31 2018 (review date August 2018)
Synthetic Biology Internal Review
Proposals are evaluated by at least three external reviewers in a process known as Synthetic Biology Internal Review (SBIR). SBIR encourages investigators to extensively consider broader aspects of their research (e.g., biosafety, biosecurity, bio-containment and environmental issues) to evaluate both positive and negative impacts and to propose strategies to mitigate concerns. If issues are not sufficiently addressed, users will be asked to modify their proposal. If issues are not resolved, the proposal may be rejected. SBIR generally takes three weeks.
Investigators should not merely write “None” or “All research will be conducted in a safe manner according to Federal regulations” in the broader implications statement, as this will lead to requests for proposal modifications, incurring delays of three weeks or longer.
Investigators must explicitly state whether their proposed research would:
- Demonstrate how to make a vaccine ineffective
- Confer resistance to antibiotics or antiviral agents
- Enhance a pathogen’s virulence or make a non-virulent microbe virulent
- Increase transmissibility of a pathogen
- Alter the host range of a pathogen
- Enable a pathogen’s ability to evade diagnostic or detection modalities
- Enable the weaponization of a biological agent or toxin