20Nov18000001_A02-L2_S2
Aplysina aerophoba
Protocols: Individuals of the sponge Aplysina aerophoba were collected by scuba diving in May 2019 at the Mediterranean coast of Spain (42.1145863 N, 3.167167 E), at a depth between 3 to 10 m. After collection, animals were transported to the Experimental Aquaria Zone (ZAE) at the Institute of Marine Science (ICM-CSIC) in Barcelona (Spain). There, each sponge individual was divided into 4 specimens with its own osculum. Each specimen was placed into individual aquaria (6 L). All the aquaria were maintained in a flow-through system with direct intake of seawater. After 5-day acclimation period, the specimens from each sponge individual were randomly assigned to either (i) control temperature or (ii) heatwave conditions. See details in treatment protocol. In the last day of experiment, half of the specimens in each temperature was challenged with Escherichia coli-derived lipopolysaccharide or with sterile artificial seawater. See detaisl in treatment protocol. 3h after exposure, samples were rinsed with sterile artificial seawater, fixed in RNAlaterTM (Invitrogen, Thermo Fisher Scientific, Germany) at 4 °C overnight. Samples were then stored at -80 °C until RNA extraction. After acclimation, sponges were maintained at either 19ºC in control conditions and 26ºC in heatwave conditions. Temperature was gradually increase (approx. 1ºC per day). Once the target temperatures were reached, sponges were maintained in these conditions for 18 days. At day 18, in each temperature condition, specimens were injected with either LPS (1mg/mL in filtered artificial seawater, source: _Escherichia coli_ O55:B5, Sigma, L2880) or sham (sterile filtered artificial seawater). Samples right at the point of injection were collected 3h post-injection. In 3 out of the 10 specimens in hot treatment, necrotic areas covered by a white film were visible, whereas nearby tissue looked intact. For those sponges, we collected two additional samples: one sample from the healthy tissue next to the necrotic area (“HT-c”) and one sample from the necrotic area (“HT-sick”) To extract total RNA, ca. 30 mg (wet weight) of sample was homogenized in 1% ß-mercaptoethanol in RLTplus buffer (Qiagen, Germany) by using the PowerLyzerTM24 (Mobio, Qiagen, Germany). Total RNA was extracted with RNeasy kit (Qiagen, Germany), treated with DNase (Ambion, Germany), and purity, quality and quantity was checked in Nanodrop 2000c Spectrophotometer (PeoLab, Germany), Experion™ electrophoresis station (Bio-Rad, Hercules, CA) and Qubit 2.0 (Life Technologies, Carlsband, CA). The absence of DNA was further checked by PCR amplification of eukaryotic 18S rRNA gene (Sigma-Aldrich, Germany: forward primer - ACCTGGTTGATCCTGCCAG, reverse primer - AATTATAATACGACTCACTATAGATTCYGCAGGTTCACCTAC; Stewart et al. 2010). Total RNA that was DNA-free and presented RIN>8 was submitted to the Competence Center for Genomic Analysis (CCGA), Kiel, Germany, for library construction and sequencing Library construction was performed with TruSeq stranded mRNA library prep kit,( Illumina, Inc., USA) with a polyA enrichment step at the Competence Center for Genomic Analysis (CCGA), Kiel, Germany.
ENA first public
2026-07-06
INSDC center name
Institute of Marine Research, Spanish National Research Council (IIM-CSIC)
INSDC status
public
Submitter Id
E-MTAB-16081:20Nov18000001_A02-L2_S2
broker name
ArrayExpress
collection date
not collected
disease
healthy
environmental stress
control
geographic location (country and/or sea)
not collected
individual
14
isolate
not applicable
organism part
whole organism
sample name
E-MTAB-16081:20Nov18000001_A02-L2_S2
scientific_name
Aplysina aerophoba
technical replicate
1