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Proteomic profiles of benthopelagic deep-sea copepods

Citation

Peters, Janna; Renz, Jasmin (2021), Proteomic profiles of benthopelagic deep-sea copepods, Dryad, Dataset, https://doi.org/10.5061/dryad.8cz8w9gpx

Abstract

In this study we used in parallel morphological, genetic, and proteomic charac­teristics of specimens of calanoid copepods from the abyssal South Atlantic to test if proteomic fingerprinting by matrix-assisted can accelerate estimating biodiversity. We cross-validated the respective molecular discrimi­nation methods with morphological identifications to establish COI and proteomic reference libraries. This data set includes all information on MALDI-TOF.

Methods

Calanoid copepods were collected in the benthopelagic boundary layer in the South Atlantic Ocean (14°58.91’ S, 29°56.48’ W) at a depth of 5139 m using an epibenthic sledge. On board, the samples were immediately fixed in 96% pure undenatured ethanol. Ethanol was exchanged within 24 hours of sampling and samples were constantly cooled for molecular analyses. Copepods were identified to morphospecies where possible (see reference table). In total 259 copepods were cut in half for further molecular analysis to allow for concurrent measurement of molecular genetic analysis and proteomic fingerprinting from the same specimens. Proteomic mass spectra were established using only the cephalosome of the individuals (except for individuals > 4 mm, where only the anterior part of the cephalosome was taken for analysis). The copepod tissue was quickly dried at room temperature. Depending on sample size 5-10 µl matrix solution (α-Cyano-4-hydroxycinnamic acid as saturated solution in 50% acetonitrile, 47.5% LC-MS grade water, and 2.5% trifluoroacetic acid) was added. After at least 10 min extraction 1.2 µl of each sample was added onto the target plate. Protein mass spectra were measured from 2 to 20 kD using a linear-mode MALDI-TOF System (Microflex LT/SH, Bruker Daltonics). Peak intensities were analyzed during random measurement in the range between 2 and 10 kDa using a centroid peak detection algorithm, a signal to noise threshold of 2 and a minimum intensity threshold of 400 with a peak resolution higher than 400 for mass spectra evaluation. Proteins/Oligonucleotide method was employed for fuzzy control with a maximal resolution 10 times above the threshold. For each sample 240 satisfactory shots were summed up. 

Spectra were analyzed using the packages MALDIquant and MALDIquantForeign for the R 3.6.2 statistical programming environment. Peaks were identified using a signal to noise ratio (SNR) of 7 after square-root transformation, Savitzky-Golay smoothing, baseline removal (SNIP-algorithm) and normalization (TIC) of spectra. Peaks were repeatedly binned until the intensity matrix reached a stable peak number (tolerance 0.002, strict approach) and missing values were interpolated from the corresponding spectrum. All peaks with a SNR <1.75 were assumed to be below detection limit and set to zero in the final peak matrix.

We provide here:  

  1. Raw files of the MALDI-TOF measurements (as zip file).
  2. Peak Matrix (intensity per m/z for all samples as csv file).
  3. Reference table (including sample ID for MALDI, identified morphotype name, sample ID for COI barcode as csv).
  4. R Code (with code to produce peak matrix from raw measurements).

 

Funding

DFG initiative 1991 , Award: RE 2808/3-1/2

DFG initiative 1991, Award: RE 2808/3-1/2