16S gene of bacterial communities: phycosphere amplicon data
To characterize bacterial community composition of the phycosphere among treatments via 16S rRNA amplicon sequencing, we collected aliquots of our bacterial filtrates that we had used for inoculation onto 0.22 m nitrocellulose filters, snap froze them in liquid nitrogen, and stored them at -80C. Filters were then thawed and incubated for 60 minutes at 56C in 30 L proteinase K, 100 L of ATL tissue lysis buffer and 300 L AL lysis buffer sourced from Qiagen. Cells were then lysed by vortexing for 10 minutes and DNA was extracted and purified using a DNeasy Blood and Tissue Kit (Qiagen, Hilden, Germany). Following the Earth Microbiome Project (https://earthmicrobiome.org/) standardized protocols, we targeted the V4 region of the 16S rRNA gene using the 515f/806r primer pair. Amplicon products were multiplexed with a unique 12 bp sequence per sample and then combined in an equal amount creating the final amplicon pool. Amplicons were then cleaned using the Qiagen UltraClean PCR Clean-Up Kit following manufacturers instructions (Qiagen, Hilden, Germany). The final pool of 16S rRNA amplicon products were sequenced on an Illumina MiSeq instrument set to read a paired end 150 bp cycle at the University of California San Diego Genomics Core (San Diego, California, United States).