In this work, samples for DNA collection from waters around an Acropora pulchra colony were taken in April 25th, 2018 (12:00 local time) at the Tema’e beach reef of Mo’orea (French Polynesia).
More...In this work, samples for DNA collection from waters around an Acropora pulchra colony were taken in April 25th, 2018 (12:00 local time) at the Tema’e beach reef of Mo’orea (French Polynesia). Water samples were withdrawn from three different local sites around the colony, at a depth of ∼1 m: (IN) between the living branches, ∼1 cm next to the polyps; (AL) deeper between the branches, ∼1 cm next to the coral skeleton colonized by turf algae; and (OUT) 2 meters down current from the patch over a sandy bottom ∼2 m deep. At that sampling point, 0.8 L (IN and AL) and 2.5 L (OUT) seawater were sampled and were prefiltered through 200-μm mesh. The microbial biomass was then collected on 0.2 μm pore-size, 47 mm diameter polycarbonate filters using a peristaltic pump. The filters were flash frozen in liquid N2 and stored at -80°C. Total DNA was extracted using the phenol-chloroform protocol as described in Massana et al. (1997). Prokaryotic diversity was determined by amplicon sequencing of the V4/V5 regions of the 16S rDNA genes using the Illumina MiSeq platform and paired-end reads (2 x 250 bp). PCR amplifications were done using the prokaryotic universal primers 515F-Y (5'- GTGYCAGCMGCCGCGGTAA-3’) and 926R (5’-CCGYCAATTYMTTTRAGTTT-3’) (Parada et al. 2016). All samples were sequenced at the Research and Testing Laboratories (RTL, Lubbock, TX, USA).
References:
Massana, R., Murray, A. E., Preston, C. M., and DeLong, E. 1302 F. (1997). Vertical distribution and phylogenetic characterization of marine planktonic Archaea in the Santa Barbara Channel. Appl. Environ. Microbiol. 63, 50–56.
Parada, A. E., Needham, D. M., and Fuhrman, J. A. (2016). Every base matters: assessing small subunit rRNA primers for marine microbiomes with mock communities, time series and global field samples: Primers for marine microbiome studies. Environ. Microbiol. 18, 1403–1414.
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