JOHN TEEM SENIOR SCIENTIFIC PROGRAM MANAGER.
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1 JOHN TEEM SENIOR SCIENTIFIC PROGRAM MANAGER
2 Synthetic biology is the design and construction of new biological entities such as enzymes, genetic circuits, and cells or the redesign of existing biological system. What makes it different from traditional biotech? Instead of introducing single genes, entire genetic pathways are introduced into cells. The genes of the pathway are modified to allow them to be assembled together as standardized parts.
3 Overview of Topics Presented The concept of interchangeable genetic parts igem (International Genetically Engineered Machine) competition Assembling pathway genes to transfer From plants into yeast From bacteria into plants From plants into plants
4 From a Synthetic Biology Perspective, Genes are Like Components Parts in a Circuit Board Promoter Terminator Coding Region
5 From a Synthetic Biology Perspective, Genes are Like Components Parts in a Circuit Board Promoter Terminator Coding Region Transcriptors Genes that function like Transisitors - Drew Endy, Stanford University
6 From a Synthetic Biology Perspective, Genes are Like Components Parts in a Circuit Board Promoter Terminator Coding Region New biological devices can be built by combing parts.
7 The genes must be recoded to remove regulatory features that may have been present in their original form. Potential problems may result from changes in flux through the pathway resulting from the removal of native regulatory features. Additionally, specific proteins in a pathway may be affected by their cellular location and require adjustments in targeting.
8 Signal Transduction Pathways Can Modified to Alter the Input Signal and Output Effect Bound attractant (sugars, amino acids, etc) Bound attractant (arsenic, estrogen, etc) Nutrient Metals, Hormones MCP MCP CHE Chemotaxis Signal Transduction CHE Chemotaxis Signal Transduction Smooth Swimming Reporter Gene Smooth Swimming Reporter Gene Expression
9 For over 10 years, igem has been encouraging students to work together to solve real-world challenges by building genetically engineered biological systems with standard, interchangeable parts. Student teams design, build and test their projects over the summer and gather to present their work and compete at the annual Jamboree. Participation in igem empowers teams to manage their own projects, advocate for their research and secure funding. Teams are also challenged to actively consider and address the safety, security and environmental implications of their work.
10 1. Design a system of genes to express in bacteria for a useful purpose (Example: Biosensor for ethylene to monitor levels in ripening fruit) 2. Search the Parts Database to find the needed component Parts 3. Order a Distribution kit with the Parts 4. Assemble the Parts using by ligation and transformation (3A assembly) 5. Test the new system and report results 6. Attend the Jamboree 7. Submit the new assembled constructs back to the repository for others to use. #Parts 2006 = = 20,000
11
12 Problem: Fertilizers are need in large amounts to provide a source of nitrogen to crop plants. Fertilizers are expensive and lead to environmental runoff problems (algal blooms). Nitrogen-fixing bacteria can supply nitrogen in some plants (legumes) that are able to form root nodules through a symbiotic association. Synthetic Biology Solution: Can nitrogen fixation genes from symbiotic nitrogenfixing bacteria be expressed directly in crop plants so that they supply their own need for fixed nitrogen?
13 Refactoring the nitrogen fixation genes from Azotobacter vinelandii Nitrogenase is unstable in the presence of O2, so it must be assembled in mitochondria in yeast. Nitrogenase subunits assembled into tetramers within mitochondria, but purified nitrogenase enzyme lacked activity.
14 Problem: Fertilizers are need in large amounts to provide a source of nitrogen to crop plants. Fertilizers are expensive and lead to environmental runoff problems (algal blooms). Nitrogen-fixing bacteria can supply nitrogen in some plants (legumes) that are able to form root nodules through a symbiotic association. Synthetic Biology Solution: Can genes from legumes needed for nodule formation be transferred to cereals so that cereals could form a symbiotic association with nitrogenfixing bacterial?
15 To Completely Reproduce the Nitrogen Symbiosis in Cereals Four Co-ordinated Genetic Programs Would Have to be Introduced into Cereals 1. Recognition of Nod factors (Nodule forming factors) 2. Organogenesis of the root nodule 3. Bacterial infection 4. Establishment of a suitable environment for nitrogenase activity inside the nodule Legume Genes Identified NFR1, NFR5, LYS3 NIN ERN1
16 The first step in nodule formation in legumes is the plant responding to Nod factor (produced by rhizobia, the nitrogen-fixing symbiotic bacteria). Nod factors, the lipochitooligosaccharide (or LCO) signals of rhizobia, are structurally very similar to those mediating signalling during the mycorrhizal symbiosis, known as Myc factors. Just like Nod factors, Myc factor perception leads to the activation of a common symbiosis signaling (SYM) pathway. Legumes Nod factors NFR1, NFR5, LYS3 SYM Pathway Cereals Myc factors SYM Pathway Engineering nodulation signalling would therefore be engineering the perception of Nod factors to activate the SYM pathway and engineering the outputs of this pathway for nodulation-specific gene expression. Nodulation-specific gene expression NIN, ERN1 Mycorrhizal-specific gene expression
17 The first step in nodule formation in legumes is the plant responding to Nod factor (produced by rhizobia, the nitrogen-fixing symbiotic bacteria). Nod factors, the lipochitooligosaccharide (or LCO) signals of rhizobia, are structurally very similar to those mediating signalling during the mycorrhizal symbiosis, known as Myc factors. Just like Nod factors, Myc factor perception leads to the activation of a common symbiosis signaling (SYM) pathway. Legumes Nod factors NFR1, NFR5, LYS3 SYM Pathway Cereals Nod factors NFR1, NFR5, LYS3 SYM Pathway Engineering nodulation signalling would therefore be engineering the perception of Nod factors to activate the SYM pathway and engineering the outputs of this pathway for nodulation-specific gene expression. Nodulation-specific gene expression NIN, ERN1 Nodulation-specific gene expression NIN, ERN1
18 The first step in nodule formation in legumes is the plant responding to Nod factor (produced by rhizobia, the nitrogen-fixing symbiotic bacteria). Legumes Cereals Nod factors Nod factors # genes? It is not yet clear how many genes will be needed, at both ends of the SYM pathway, to allow cereals respond to rhizobia with nodule formation. # genes? NFR1, NFR5, LYS3 SYM Pathway Nodulation-specific gene expression NIN, ERN1 NFR1, NFR5, LYS3 SYM Pathway Nodulation-specific gene expression NIN, ERN1
19 Synthetic Biology for Nitrogen Fixation in Cereals Expression of bacterial nitrogen fixation genes in plants is not yet feasible, and will probably require troubleshooting using yeast as an intermediate model eukaryotic system. Engineering cereals to form symbiotic association with nitrogen-fixing bacteria is also not yet feasible, and will require further understanding of the role of legume plant genes involved in Nod factor recognition and nodule formation.
20 Synthetic Biology for Agricultural Biotechnology Will require efforts analogous to igem for designing new genetic circuits in plants. Will require the development of surrogate systems (like yeast) to facilitate the transfer of metabolic pathways.
21 Are existing risk analysis frameworks sufficient to address the risks poses by Synthetic Biology? Risk assessments must considered on a case by case basis, taking into account the specific properties of the GE organism in relation to the receiving environment. Synthetic Biology makes use of the same genes that comprise traditional GE plants, so the existing methodology should be sufficient to address risks associated with their use as component parts. As with traditional GE plants, the use of problem formulation will be an essential part of the risk assessment process for GE organisms incorporating Synthetic Biology.
22 Thank You!
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