5-ALA is a widely used biopesticide, which can effectively improve the resistance of crops to biotic and abiotic stresses. Promoting high yields of 5-ALA is a priority in agricultural research. However, in the current research on high-yielding 5-ALA, there is still a lack of a high-throughput screening method for high-yielding strains. While striving to achieve high yield of 5-ALA, we are also actively exploring and hoping to establish a high-throughput high-yield strain screening method with the help of droplet microfluidic technology.
In the experimental exploration of droplet microfluidic high-throughput screening, we explored a stable droplet generation system suitable for E. coli encapsulation after preliminary experimental exploration, and successfully used the difference in strain biomass to complete the screening of the high-yield 5-ALA system, and the fermentation results were verified. However, due to the low accuracy and large error of the absorbance screening method,[1]this is also reflected in our fermentation validation results. Although it is possible to screen for high-yielding strains, their numbers are small and not ideal.
We found that cells can produce different amounts of metabolites at different stages of growth, so sorting can be performed based on the fluorescence signal produced by the metabolites. Cell analysis based on fluorescence signal is the most commonly used method, which does not require the addition of other reagents, and has the characteristics of simplicity, high sensitivity, high throughput, and wide range of applications.
However, due to the limitation of time and working conditions, we are not able to further explore the fluorescence intensity screening with higher sensitivity, so we hope to explore the fluorescence screening of droplet microfluidics more deeply in the later work.
We hope that the proportional relationship between protoporphyrin and the target product 5-ALA can be effectively verified through experiments, and with the help of biological companies, the screening of high-yielding strains of 5-ALA can be completed by using the corresponding fluorescence screening equipment, so as to establish a more accurate and efficient system strain screening method.
We also hope that through our experimental exploration process, we can give other iGEM teams a new way of thinking. This improves the efficiency and accuracy of traditional screening, making our experiments faster, more accurate, and more efficient in the future.