Anemia affects one third of the world's population and is mostly due to insufficient iron intakes. Anemia and similar mineral deficiency diseases are primarily widespread in developing countries like India, partly resulting from the local diet that is mainly made up of cereal grains and seeds such as rice . In these types of food, iron bioavailability is substantially reduced by the presence of phytic acid (C₆H₁₈O₂₄P₆), which chelates minerals and forms insoluble salts which precludes their absorption in the gastrointestinal tract.

Current research on increasing the bioavailability of iron or zinc involves the bioengineering of crop plants which not only poses challenges in terms of the production of efficient genetically modified crops but also requires extensive research for drawing any conclusions on strain sustainability.

We propose an alternative strategy that focuses on the bioengineering of microorganisms involved in the fermentation of idli, a dish widely used as primary food source throughout much of India. Indeed, the lab model organism Saccharomyces cerevisiae is a strain present in the idli microbiome that naturally produces phytases. Phytases are enzymes that are able to hydrolyze phytic acid even when complexed with minerals, resulting in a greater mineral bioavailability.

However the production of phytases in Saccharomyces cerevisiae is down-regulated by two genes: PHO80, present on chromosome 15 and PHO85, found on chromosome 16. The knockout of these genes would likely increase the yield of phytase production and therefore increase the general bioavailability of minerals in fermentation-based dishes such as idli.

We used a strain of Saccharomyces cerevisiae(SK1), a gift of the INSERM unit U1001. We designed a set of primers to delete the two phytase synthesis repressor genes PHO80 and PHO85 on chromosomes 15 and 16. We amplified the geneticin resitance gene(G418) from the pSB1C3 plasmid with primers designed to add 2 homology region in order to insert the resistance gene inside the genes we wanted to delete.

After the successful transformation and selection (fig5), we measured whether the level of phytase production was increased. This was done through the use of a quantification colorimetry kit, which measures the OD655 to determine how much phytic acid is found in different growth conditions. In order to do that, we used the kit to quantify the levels of phytic acid in rice, fermented rice and idli at different stages of it's preparation using our different strains.

In Table 1, we can see the typical data that we obtain after usage of the kit. The values are very low in comparison with what is expected for such food samples. (for rice is is supposed to be equal to 6 mg.g^-1, and here we observe a concentration of 5 µg.g^-1, 10^-3 less).

The figure 3 is showing the variation of OD655nm before and after growth with different strains/food samples.The results are not showing any significant decrease in concentration after growth or even between samples that were expected to present different levels of free phosphorus.

The results for the comparative growth experiment are showing no significant decrease between the growth speed of the yeasts before and after deletion of the gene(fig4).