Difference between revisions of "Team:IIT-Madras/Description"

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<li>Lignins are important in the formation of cell walls, especially in wood and bark, because they lend rigidity and do not rot easily. Lignin is rich in aromatic compounds which are degraded only by few organisms. Lot of work has been done on degrading aromatic compounds using Pseudomonas as chassis but even it has its own limitations. So, there is a pressing need to use other chassis that can complement the work. Acinetobacter baylyi ADP1 fits in exactly as due to its ability to degrade aromatic compounds and lignin-derived monomers.</li>
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<li><i>Acinetobacter baylyi</i> ADP1 possesses catabolic pathways to degrade aromatic compounds. These pathways can be re-engineered to produce biofuels from Lignin monomers which is abundant in plant biomass. It can also be useful to produce bio-surfactants and lubircants such as Wax-Ester (Kannisto et al. 2016, Journal of Industrial Microbiology and Biotechnology). These pathways are not present in many organisms. Some work has been done on degrading aromatic compounds using Pseudomonas as chassis but even it has its own limitations. Acinetobacter baylyi ADP1 complements abilities and applications of Pseudomonas. </li>
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<li>For routine model organisms like E. coli and L. lactis., they are not naturally competent and so we need to artificially make them competent. We found many ‘Naturally Competent’ microorganisms. However, most of them are pathogenic and known to cause harm to humans. Hence, you cannot work with them in Biosafety level 1 lab. For example Streptococcus pneumoniae, Neisseria gonorrhoeae, Bacillus subtilis and Haemophilus influenzae.
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<li>For routine model organisms like E. coli and L. lactis., they are not naturally competent and so we need to artificially make them competent. We found many ‘Naturally Competent’ microorganisms. However, most of them are pathogenic and known to cause harm to humans. Hence, it is not possible work with them in Biosafety level 1 lab. For example Streptococcus pneumoniae, Neisseria gonorrhoeae, Bacillus subtilis and Haemophilus influenzae.
 
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Revision as of 13:45, 9 November 2018

iGEM Collaborations Page

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Team: IIT-Madras/ADaPtat1on

Description

ADaPtat1on

Motivations:

  1. Acinetobacter baylyi ADP1 possesses catabolic pathways to degrade aromatic compounds. These pathways can be re-engineered to produce biofuels from Lignin monomers which is abundant in plant biomass. It can also be useful to produce bio-surfactants and lubircants such as Wax-Ester (Kannisto et al. 2016, Journal of Industrial Microbiology and Biotechnology). These pathways are not present in many organisms. Some work has been done on degrading aromatic compounds using Pseudomonas as chassis but even it has its own limitations. Acinetobacter baylyi ADP1 complements abilities and applications of Pseudomonas.
  2. For routine model organisms like E. coli and L. lactis., they are not naturally competent and so we need to artificially make them competent. We found many ‘Naturally Competent’ microorganisms. However, most of them are pathogenic and known to cause harm to humans. Hence, it is not possible work with them in Biosafety level 1 lab. For example Streptococcus pneumoniae, Neisseria gonorrhoeae, Bacillus subtilis and Haemophilus influenzae.



Background:

We found from the literature that Acinetobacter baylyi ADP1 to be a good chassis for our project because of its ability to degrade aromatic compounds and it is naturally competent. It is also non-pathogenic and belongs to risk group 1[1]. Also, recent work has been done to produce wax ester in Acinetobacter baylyi ADP1 [2]. Strains of Acinetobacter baylyi have been constructed that can utilize Gluconate and Glucose better than the existing strain [3]. However, a big shortcoming of using this organism is that it does not many tools for gene manipulation. For example, usually, T5 and T7 are the only two standard promoters used in engineering this organism.



Project:

Our objective is to create a T5 based synthetic promoter library for Acinetobacter baylyi ADP1. For this purpose, we also needed fluorescent reporter protein as we intended to measure the strength of promoter using fluorometry experiment. We approached GenScript for codon optimized GFP for Acinetobacter baylyi ADP1, however, they did not have the codon usage table data for Acinetobacter baylyi. .

We made a free use online tool called CUTE(codon usage table easy) that can generate Codon usage table by taking into consideration the protein-coding annotation. This tool can be used for any other organism whose coding regions are annotated. Cute can be found on the cute.chassidex.org
We generated the codon usage table as the protein annotation of Acinetobacter baylyi ADP1 is available on the NCBI website. Using this table, we codon optimized GFP (which was codon optimized previously for E. coli).
Next, we generated the T5 promoter-based library for Acinetobacter baylyi ADP1. Since these promoters are T5 based, they might potentially also work in other gram-negative organisms like E. coli strains, Cornybacterium etc.


References:

  1. https://www.dsmz.de/catalogues/details/culture/DSM-24193.html?tx_dsmzresources_pi5%5BreturnPid%5D=304
  2. Suvi Santala, Elena Efimova, Perttu Koskinen, Matti Tapani Karp, and Ville Santala ACS Synthetic Biology 2014 3 (3), 145-151 DOI: 10.1021/sb4000788
  3. Kannisto, Matti et al. “Metabolic Engineering of Acinetobacter Baylyi ADP1 for Improved Growth on Gluconate and Glucose.” Ed. S.-J. Liu. Applied and Environmental Microbiology 80.22 (2014): 7021–7027. PMC. Web. 17 Oct. 2018

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