e5 test samples for metabolomics

We are sending test samples to the UW metabolomics factility to decide if we want to send out the full timeseries for metabolomics. The UW facility has never done coral samples before so they are letting us send four free test samples to see if they will work. See github issue here.

I’m using the following samples for tests. These are samples that are the same species as our sequencing samples but are not going to be used for sequencing/metabolomics.

• POC-68 TP1
• POR-70 TP2
• ACR-140 TP2
• ACR-165 TP1

Equipment & materials

• Ice / dry ice
• Weigh boat
• Scale
• 10x PBS
• Plastic pestles (can fit in 1.5 mL tube)
• 1.5 mL tubes
• 50 mL tubes
• Centrifuge
• Pipettes + tips
• Airbrush

Protocols

IMPORTANT: Keep everything as COLD as possible; keep tubes on ice at all times when not in centrifuge.

IMPORTANT: When airbrushing, clean materials with 10% bleach, 70% ethanol and DI water in between samples.

• Make 10x PBS in autoclaved container and keep on ice
• Thaw fragment on ice and weigh mass of fragment
• Airbrush with ice cold 10X PBS
• Pour into 50 mL tube and record slurry volume
• Aliquot 1 mL of slurry into a 1.5 mL tube
• Homogenize on ice using plastic pestle in tube to avoid lipid oxidation (do not use immersion homogenizer)
• Separate fractions by centrifuging at 3,000g for 5 min at 4°C
• Move supernatant to new tube and try to keep/scrape lipid films to keep with the supernatant
• Vortex briefly and centrifuge again at 3,000g for 5 min at 4°C
• Move supernatant to new tube, this is the host tissue; keep any lipid films as much as possible
• Resuspend the symbiont pellet with 1 mL of 10x PBS
• Return tubes to -80°C
• Air dry airbrushed fragments overnight
• Weigh mass of airbrushed fragment and record difference in weight
• Ship to UW on dry ice directly to facility (this will be done on Tuesday 9/3)

Here’s the info for the test samples:

Colony_ID	Species	Timepoint	DateAirbrushed	Weight_PreAirbrush_g	Weight_PostAirbrush_g	Weight_Difference_g	HomogenateVolume_mL
POC-68	Pocillopora tuahiniensis	TP1	20240829	7.4847	6.337	1.1477	15
POR-70	Porites evermanni	TP2	20240829	0.6733	0.3881	0.2852	12
ACR-140	Acropora pulchra	TP2	20240829	0.6726	0.5975	0.0751	9
ACR-165	Acropora pulchra	TP1	20240829	1.2964	1.0039	0.2925	12

Here’s what the fragments looked like post-airbrushing: