The Chlamydomonas CC-4533 strain was grown in TAP medium in a 20-liter container under 100 μmol photons m-2 s-1 light (measured at the periphery) to a density of 1-1.5 × 10^6 cells/mL. Cells were collected by centrifugation at 300-1000 x g for 4 min. Pellets were washed once with 25 mL TAP supplemented with 40mM sucrose, and then resuspended in TAP supplemented with 40 mM sucrose at 2 x 10^8 cells/mL. 250 µL cell suspension was then aliquoted into each electroporation cuvette (1652088, Bio-Rad) and incubated at 16°C for 5-30 min. For each cuvettee, 5 μL of cassette CIB1 DNA at 5 ng/μL was added to the cell suspension and mixed by pipetting. Electroporation was immediately performed with the procedure described before (Zhang, Patena et al. 2014). After electroporation, cells from each cuvette were diluted into 8 mL TAP supplemented with 40 mM sucrose and shaken gently in dark for 6 h. After incubation, cells were plated on TAP containing 20 µg/mL paromomycin (800 µL per plate) and incubated in dark for approximately two weeks before colony picking.