Bacterial metagenome of Durusdinium trenchii strain CCMP3408 lab culture
Microbiome samples from each Symbiodiniaceae culture, as well as negative controls, were collected at about the same time of day. Only bacteria that were physically attached to the Symbiodiniaceae cell surface were sampled. DNA extraction was performed using the DNeasy PowerSoil Pro kit (Qiagen) according to the manufacturers protocol. Two replicates of a mock community, the ZymoBIOMICSMicrobial Community Standard (Zymo Research), were included as positive controls. The full 16S rRNA gene was amplified in 25 l PCR reactions. A sample of autoclaved nuclease-free water was included as a PCR negative control. Due to low yield of bacterial DNA extracts and thus PCR products, each PCR reaction was replicated six times and then pooled between all six replicates, then purified using the DNA Clean & Concentrator-5 PCR purification kit (Zymo Research) and AMPure XP beads (Agencourt AMPURE XP, Beckman Coulter). A multiplexed library (20 barcodes) was prepared using the Native Barcoding Kit 24 V14 (SQK-NBD114.24) and loaded onto separate R10.4.1 flow cells (FLO-MIN114). Sequencing was performed on a MinION Mk1C controlled by MinKNOW v 23.07.12 on an iMac (High Sierra Version 10.13.6, Intel Core i7, 32GB, 4 cores) connected by USB, with real-time basecalling.