A green micro-algal growth media modified for use as a stringent minimal media for bacteria. v1
Authors: Mitra Mautusi 
Published: June 2020

Introduction: Chlamydomonas reinhardtii, a green micro-alga can be grown at the lab heterotrophically or photo-heterotrophically at room temperature using the Tris-Phosphate-Acetate (TAP) medium which contains 0.1 % acetate (acetic acid) as the sole carbon source. Standard TAP medium recipe can be found at the website of Chlamydomonas Resource Center: https://www.chlamycollection.org/methods/media-recipes/tap-and-tris-minimal/. Hutner’s trace element solution is an ingredient in the TAP medium. Hutner’s trace element recipe can be found at https://www.chlamycollection.org/methods/media-recipes/hutners-trace-elements/. M9 medium is the standard minimal medium for growing bacteria (http://www.thelabrat.com/protocols/m9minimal.shtml). When grown in TAP medium, Chlamydomonas can utilize the exogenous acetate in the medium to make net biosynthesis of sugar using the Glyoxylate cycle and can grow either heterotrophically or photo-heterotrophically without being dependent on photosynthesis for glucose biosynthesis. Higher green plants have the glyoxylate cycle which they use to make sugar from internal pool of acetyl CoA but cannot take up exogenous acetate like Chlamydomonas. In our lab we use a slightly modified TAP medium recipe which has a final concentration of phosphate, nitrogen, magnesium and calcium approximately 10-fold, 2-fold, 2-fold and 2-fold higher than that in the TAP recipe described on the Chlamydomonas Resource Center website, respectively. Final concentrations of acetate and Hutner’s trace elements are same in both TAP recipes. Attached Table 1 compares the chemical ingredients in our lab’s TAP recipe (full recipe can be found on protol.io) with that present in the standard M9 medium described at http://www.thelabrat.com/protocols/m9minimal.shtml. M9 medium has a final concentration of phosphate, nitrogen, magnesium and carbon, approximately 70-fold, 2.5-fold, 4-fold and 5-fold higher than that present in our lab’s TAP medium, respectively. Additionally, M9 contains 0.05% salt (8.56 mM) (Table 1). TAP has additional trace elements like iron, zinc, copper, manganese, cobalt, boron and molybdenum which are components in the Hutner’s trace element solution (Table1). In summary, TAP medium is chemical composition wise, a more stringent minimal medium than the M9 medium used by microbiologists. Applications: There are many bacteria that can also utilize acetate as a carbon source. Some of these bacteria also have glyoxylate cycle like Chlamydomonas and green plants. Hence, TAP medium can be used to isolate acetate-requiring bacteria. If acetate is removed from the TAP medium, it becomes the TP medium which lacks a carbon source and will not allow Chlamydomonas or other acetate-requiring bacteria to grow. The 0.1 % acetate in the TAP medium can be substituted with alternative carbon sources (e.g. glucose, sucrose, lactose, hydrocarbons, aromatic compounds and polyhydroxyalkanoates etc.) to test physiological abilities of candidate bacteria to use the tested chemicals as the sole alternative carbon and energy source.