Symbiodiniaceae spp. extracted from the sea anemone Exaiptasia pallida during heat stress experiments
PCRs were performed in triplicates using the primers SYM_VAR_5.82S2 (5 TCGTCGGCAGCGTCAGATGTGTATAAGAGACAG-GAATTGCAGAACTCCGTGAACC 3) and SYM_VAR_REV (5GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAG-CGGGTTCWCTTGTYTGACTTCATGC 3). Each PCR reaction was run with Qiagen Multiplex PCR Kit (Qiagen, Germany) and 10 M primers in a final reaction volume of 15 L. Thermal cycling conditions of 15 min at 95 C, followed by 30 cycles of 30 s at 95 C, 90 s at 56 C and 30 s at 72 C, with a final extension step of 10 min at 72 C were used for amplification. For each sample, PCR products were run on a 1 % agarose gel electrophoresis, pooled and cleaned using ExoProStar 1-step (GE Healthcare, Little Chalfont, UK). Indexing was then performed using the Nextera XT Index Kit (Illumina, CA, USA) according to the manufacturers instructions and followed by sample normalization and final library pooling. A SequalPrep Normalization Plate Kit (Invitrogen, Thermo Fisher Scientific, USA) was used to do the normalization, avoiding the more labor-intensive process of quantifying and aliquoting each individual sample. The final pooled library was quantified on a BioAnalyzer (Agilent Technologies, CA, USA) and sequenced at 7 pM with 20 % phiX on the Illumina MiSeq, 2  300 bp end version 3 chemistry according to the manufacturer's specifications at the Bioscience Core Lab in KAUST, Saudi Arabia