Our project focused on developing methods to tame non-model organisms and develop them for genetic manipulation. We chose for this project Cyanobacterium and Rhizobium because we foresee their successful applications in the fields of biotechnology. Cyanobacteria can become a producer of biofuel driven by light energy and Rhizobium can cultivate the soil and control the spread of genetically modified crops.
We succeeded in growing the strains and developing a robust assay for testing growth of non-model organisms in various media. We were informed by literature and work previously done in growing these particular microbes.
We were able to transform our non-model organisms and express our heterogenous DNA. Transformation is an integral part of synthetic biology and so we spent significant effort trying multiple protocols until we arrived at one with acceptable efficiency
After transformation, selection was a key step. By modifying existing protocols testing for antibiotic resistance, we were able to develop a robust assay for antibiotic resistance and susceptibility in non-model organisms. This was a key step in pushing forward in taming non-model organisms.
Multiplex automated genome engineering (MAGE) and recombineering are genetic manipulations techniques now used in E. coli, but that would make ideal techniques for use in our non-model organisms. In order to establish these in our organisms we searched through the body of research currently in existence and pulled out potentially useful recombinase proteins. We developed an assay to test for the activity of our recombinases as well.
Taming the Model Organism
Establishing genetic manipulation techniques and standard practice for these non-model organisms would have been the absolute success for our project. Thus far we have determined how to grow, transform, select, and integrate important elements of genetic manipulation systems into our organisms. More importantly, we have developed assays and principles that can be applied to the taming of any non-model organism.