From 2011.igem.org

Revision as of 16:52, 25 October 2011

Informing Design

We consulted numerous experts in various fields to ensure that the design of the AuxIn system respects all relevant social, ethical and legal issues. One module of our system, Gene Guard, is a direct result of brainstorming around the issues involved in the release of genetically modified organisms (GMOs). Although we have only reached the proof of concept stage, we have put a lot of thought into how AuxIn may be implemented as a product and the legal issues that would be involved.

Ecology

Since our project involves releasing bacteria into nature, we have to consider their impact on the environment and other soil organisms. This is vital in assessing the safety of release. We did a literature review of the impact of natural auxins on the rhizosphere and consulted ecologists to find out more about the impact our GMO could have.

Effect on soil fauna

Figure 1. Dr Alex Milcu demonstrating the Ecotron experimental setup (photo courtesy of Dr Milcu).

1. Goal

Many animals above and below ground are extremely important to existing ecosystems. We want to ensure that there is no significantly detrimental effect on these animals caused by our bacteria.

2. Action

We met Dr Alexandru Milcu, an expert in above/below ground interactions at the Imperial College Silwood Park campus to discuss the impact of our project on the soil ecosystem (see Figure 1).

Dr Milcu informed us that there is no known negative effect of auxin on soil fauna. A more fibrous root network should not influence metazoans to a noticeable extent. However, earthworms take up nutrients through their coelom and may be affected by heightened auxin concentrations, although this has not been established in the literature. Earthworms act as "ecosystem engineers" and are therefore extremely important to the soil. Affecting them negatively should definitely be avoided (Dr Alexandru Milcu, oral communication).

3. Result

We devised a development plan (see the Implementation page) that our project will undergo before release would become feasible. This plan includes extensive testing of the impact of AuxIn on several important organisms, including earthworms, fish and other animals.

Effect of protozoans on our project

1. Goal

An issue with dispersing bacteria in the soil is that they may be eaten by protozoa. Grazing by protozoa has been shown to increase plant growth ^[6]. There are competing theories that this may be due to promoting auxin-producing bacteria or by increasing the nitrogen in the soil. However, it is becoming more likely that the latter theory is true (Dr Alexandru Milcu, oral communication).

2. Action

We met with Dr Milcu to discuss the impact protozoans may have on the implementation of our project. To test the impact of protozoan grazing on our bacteria, Dr Milcu suggested ensuring that protozoa be in test compost with our bacteria and then measuring auxin levels. This would add to the complexity of trials but give a more realistic measure of density and auxin secretion than by simply plating out our bacteria on agar.

3. Result

We unfortunately did not have time to test our auxin-secreting bacteria in compost. However, we have incorporated this experiment into our future implementation plan.

Effect on plant population composition

1. Goal

Monocots such as grasses typically grow fibrous root networks, while dicots tend to grow long, deep branched roots. Auxin influences dicots and monocots differently. While it induces lateral root growth and inhibits deep root growth in both types of plants, only diots are influenced negatively by exogenous auxin^[3]. Supplying auxin to the soil may therefore result in selecting against dicots and causing a predominantly monocot population. While this may be detrimental for crop usage, dense networks of roots lead to an exponential decrease in water erosion^[4]. Fibrous root networks are therefore advantageous when trying to prevent soil erosion. We want to ensure that our bacteria do not have a detrimental effect on plant population composition.

2. Action

Figure 2. Syngenta visit: Prof. Stuart Dunbar showed us the greenhouses. (Picture by Imperial College London iGEM team 2011).

We met with soil and auxin specialists at Syngenta to discuss the implications of our project (see Figure 2). Dr Torquil Fraser and Dr. John Paul Evans advised us that there are four main natural and four synthetic auxins. Synthetic auxins are very stable and persist for weeks to months in the soil and are commonly used as herbicides. Unlike synthetic auxins, natural auxins are rapidly degraded. In plants, the auxin concentration is tightly regulated by conjugation with aspartic acid and metabolic feedback loop. This is not the case with synthetic auxins because of a slight change in their molecular structure.

In addition, we discussed the issue of skewing the plant population with Dr. Milcu.

3. Result

Skewing the plant population may have negative impacts from an ecological perspective, affecting diversity, but this effect may be counterbalanced by the conservation of plant habitation in general. In addition, we are considering putting our bacteria into soil alongside plants that are able to revegetate desertified areas.

One of the great advantages of synthetic biology is the degree of control we can have on a particular system. We have introduced a sequence between RBS and promoter that facilitates exchanging promoters without affecting the coding sequence. It will therefore be possible to change the amount of IAA secreted by our bacteria to ensure that the amount produced is not inhibitory.

To test the effects of heightened auxin on different kinds of plants, Dr Milcu suggested setting up lab experiments with different plant functional groups. These would be planted in compost with an increased auxin concentration (using synthetic auxin) and observe how they respond. This experiment would be performed in the later testing stages of the project when we would be testing the impact of auxin on the environment.

PGP's and you

Plant growth promoting bacteria

The close mutualistic relationship between plants and funghi (for ex. micorhhiza) has been known for long, but plants do not only live in close synbiosis with many species of funghi but also with bacteria. Similarly to the human body, the body of a plant is also full of bacteria living by, on and in it that interact with the plant continuously. Symbiotic bacteria benefit plants in many ways[6][7]. They help to keep away parasites, take up minerals from the soil (eg nitrogen fixation), enhance stress tolerance and increase growth.

Many of them do so by producing various compounds that stimulate plants, one of which is auxin[8]. This strategy of natural bacteria in order to increase root growth is the basis of project AuxIn. Using the tools of Synthetic Biology, this root growth promoting strategy can be tailored in order to meet especially the needs of solving the problem of desertification.

Why engineer bacteria instead of plants?

The close relationship between bacteria and plants has been evolving for long time and could open a wide gate to use synthetic biological tools in order to influence plants.

There are many advantages of using this approach; the most straightforward is the fact that engineering bacteria is easier than engineering plants. Also, through the genetically engineered machine concept, it is possible to have tight control and fine tuning of the way engineered bacteria carry out their task.

Furthermore, often the plant in question can be one that is poorly characterised in terms of use in lab which could make it even harder to work with. On the other hand, a specie of bacteria can interact with many different plant species and could be adjusted to the plant specie in question. Therefore a certain type of GM bacteria (eg. Auxin producing) could be used with lots of plants all around the globe. Therefore in many cases the GM bacteria approach could be especially advantageous such as the urging problem of desertification or quick need for bioremediation in various areas.

However, crop plants are different in the terms that they are well known in all aspects and there is big effort put into improvement of existing cultivars. The improvement of drought-tolerant crop plants via genetic engineering does seem promising and there has been great progress made in the field during the past years[9]. But it should also be taken into account that the agricultural environment is different in many ways to a natural ecosystem.

References and Bibliography

[1] UNCCD (2011) Desertification: a visual synthesis. (Online) Available from: http://www.unccd.int/knowledge/docs/Desertification-EN.pdf (Accessed on 12 August, 2011).

[2] Geist H and Lambin E (2004) Dynamic causal patterns of desertification. BioScience 54: 817-829.

[3] McSteen P (2010) Auxin and monocot development. Cold Spring Harb Perspect Biol, a001479.

[4] Gyssel G and Poesen J (2003) The importance of plant root characteristics in controlling concentrated flow erosion rates. Earth surface processes and landforms 28: 371-384.

[5] Spaepe S et al. (2007) Indole-3-acetic acid in microbial and microorganism-plant signalling. FEMS Microbiol Rev 31: 425-448.

[6] Mei C, Flinn BS. The use of beneficial microbial endophytes for plant biomass and stress tolerance improvement. Recent patents on biotechnology 2010 2010-Jan;4(1): pp. 81-95.

[7] Sturz AV, Christie BR, Nowak J. Bacterial endophytes: Potential role in developing sustainable systems of crop production. Critical Reviews in Plant Sciences 2000 2000;19(1): pp. 1-30.

[8] Spaepen S, Vanderleyden J, Remans R. Indole-3-acetic acid in microbial and microorganism-plant signaling. FEMS microbiology reviews 2007 JUL 2007;31(4): pp. 425-448.

[9] Yang S, Vanderbeld B, Wan J, Huang Y. Narrowing Down the Targets: Towards Successful Genetic Engineering of Drought-Tolerant Crops. Molecular Plant 2010 MAY 2010;3(3): pp. 469-490.

[6] Krome K et al. (2009) Soil bacteria and protozoa affect branching via effects on the auxin and cytokinin balance in plants. Plant Soil 328: 191-201.

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