Dataset: Data and analysis code used to experimentally evolve representatives of four phytoplankton functional types in co-culture with a heterotrophic bacterium under either present-day or predicted future pCO2 conditions

Experimental evolution: Cultures for experimental evolution were initiated with 12.3 milliliters (mL) of media, 0.2 mL of acid or base additions for carbonate chemistry manipulation, and 0.5 mL of a previous culture. All cultures were grown at 22 degrees Celsius (°C) under approximately 75 micromoles photons per square meter per second (µmol photons m-2 s-1) in acid-washed conical-bottom glass tubes with airtight caps; with 13 mL of culture, almost no headspace existed in these tubes. All cultures except Prochlorococcus were grown on a rotating test tube wheel with illumination from the top and bottom; preliminary observations indicated that Prochlorococcus cells did not settle noticeably when grown in static test tube racks, whereas all other taxa did. Each phytoplankton clone was split into two culture lines, one maintained at 400 ppm pCO2 and the other at 800 ppm pCO2. For all strains except Synechocystis, each clone was co-cultured with a single Alteromonas clone.

Phytoplankton growth was measured every 48 hours using a Guava HT1 flow cytometer equipped with a 488 nanometer (nm) laser. Phytoplankton populations were identified by their clustering pattern on logarithmic plots of forward light scatter vs. 660 nm (chlorophyll) fluorescence. Cell densities within user-defined gates encircling the phytoplankton were calculated automatically by the Guava software. When phytoplankton cell densities crossed a cutoff value, cultures were diluted 26-fold (0.5 mL into a total volume of 13 mL) into fresh media. We targeted 108 transfers for each lineage, representing log₂26 or 4.7 generations (although we did not achieve this goal for some lineages due to repeated crashes or contamination). Samples from each lineage were cryopreserved every 25 generations and again at the end of the experiment.

Whole genome re-sequencing: Post-evolution cultures were split into 5 replicate 13 mL culture tubes, grown to the cutoff transfer cell density, and then collected by gentle vacuum filtration on 0.2 micrometer (µm) pore size polycarbonate filters, then flash frozen in liquid nitrogen and stored at -80°C. Genomic DNA was extracted from the filters using MoBio ProSoil kits, with the bead-beating step accomplished using a MP FastPrep-24 homogenizer. DNA was fragmented, ligated with Illumina adapters, and sequenced on an Illumina NextSeq500 device.