Sulfitobacter D7 and Marinobacter D6 were isolated from a coculture of E. huxleyi 379 with CAM at 7 days of growth. Bacterial populations in cocultures were stained with the live nucleic acid fluorescent marker SYTO 13 (Molecular Probes). Two distinct subpopulations were observed in CAM-treated cultures (fig. S1B) and were sorted at room temperature on the basis of green fluorescence intensity (530/30 nm) in purity mode using a BD FACSAria II cell sorter equipped with a 488-nm laser. Sorted populations were independently plated on marine agar 2216 plates (Difco) and incubated in the dark at 18°C. Sulfitobacter D7 and Marinobacter D6 were each isolated from a single colony and streaked three times from a single colony to ensure isolation of a single bacterial strain. For identification, DNA was extracted from a single colony of Sulfitobacter D7 and Marinobacter D6 using REDExtract-N-Amp Plant PCR kit (Sigma-Aldrich) according to the manufacturer’s instructions and was used as a template for PCR with general primers for bacterial 16S ribosomal RNA (rRNA): 5′-agtttgatcctggctcag-3′ (forward) and 5′-taccttgttacgacttcacccca-3′ (reverse) (58). Amplicons were paired-end sequenced using the ABI 3730 DNA Analyzer and manually assembled. Sulfitobacter D7 and Marinobacter D6 were grown in marine broth 2216 (Difco) and stored in 15% glycerol at −80°C.

Note: The content above has been extracted from a research article, so it may not display correctly.

Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.

We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.