#METABOLOMICS WORKBENCH ramkhattri_20201222_082236 DATATRACK_ID:2370 STUDY_ID:ST001645 ANALYSIS_ID:AN002691 PROJECT_ID:000000 VERSION 1 CREATED_ON January 12, 2021, 12:34 pm #PROJECT PR:PROJECT_TITLE Intraspecific variation in polar and nonpolar metabolite profiles of a PR:PROJECT_TITLE threatened Caribbean coral PR:PROJECT_TYPE intraspecific variability PR:PROJECT_SUMMARY This project aims to identify differences in metabolomic profiles among seven PR:PROJECT_SUMMARY known, unique genotypes of the threatened staghorn coral Acropora cervicornis. PR:INSTITUTE University of Florida PR:DEPARTMENT Department of Fisheries and Aquatic Sciences PR:LABORATORY Patterson Lab PR:LAST_NAME Patterson PR:FIRST_NAME Joshua PR:ADDRESS Florida Aquarium Center for Conservation, 529 Estuary Shore Lane, Apollo Beach, PR:ADDRESS FL 33572 PR:EMAIL joshpatterson@ufl.edu PR:PHONE (813) 419-4917 PR:FUNDING_SOURCE This study was funded by the University of Florida’s Southeast Center for PR:FUNDING_SOURCE Integrated Metabolomics through grant number U24DK097209 from the National PR:FUNDING_SOURCE Institute of Health’s Common Fund metabolomics program. Additional assistance PR:FUNDING_SOURCE was provided by the National High Magnetic Field Laboratory supported by PR:FUNDING_SOURCE National Science Foundation Cooperative Agreement No. DMR-1644779 and the State PR:FUNDING_SOURCE of Florida. Partial support was provided by USDA National Institute of Food and PR:FUNDING_SOURCE Agriculture HATCH Project [FLA-FOR-005902]. PR:CONTRIBUTORS Joseph A. Henrya, Kathryn E. Lohr , Ram B. Khattri, Joy Guingab-Cagmat, Matthew PR:CONTRIBUTORS E. Merritt, Timothy J. Garrett, and Joshua T. Patterson #STUDY ST:STUDY_TITLE Variability in metabolomic profiles among unique genotypes of Acropora ST:STUDY_TITLE cervicornis (part -II) ST:STUDY_TYPE intraspecific variability ST:STUDY_SUMMARY This project aims to identify differences in metabolomic profiles among seven ST:STUDY_SUMMARY known, unique genotypes of the threatened staghorn coral Acropora cervicornis. ST:INSTITUTE University of Florida ST:DEPARTMENT SECIM ST:LAST_NAME Patterson ST:FIRST_NAME Joshua ST:ADDRESS Florida Aquarium Center for Conservation, 529 Estuary Shore Lane, Apollo Beach, ST:ADDRESS FL 33572 ST:EMAIL joshpatterson@ufl.edu ST:PHONE (813) 419-4917 ST:NUM_GROUPS 7 ST:TOTAL_SUBJECTS 41 #SUBJECT SU:SUBJECT_TYPE Other organism SU:SUBJECT_SPECIES Acropora cervicornis SU:TAXONOMY_ID 6130 SU:GENOTYPE_STRAIN U77, U44, U41, U25, K1, K2, K3 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - U77_A1 Genotype:U77 RAW_FILE_NAME=U77_A1.raw SUBJECT_SAMPLE_FACTORS - U77_A2 Genotype:U77 RAW_FILE_NAME=U77_A2.raw SUBJECT_SAMPLE_FACTORS - U77_B1 Genotype:U77 RAW_FILE_NAME=U77_B1.raw SUBJECT_SAMPLE_FACTORS - U77_B2 Genotype:U77 RAW_FILE_NAME=U77_B2.raw SUBJECT_SAMPLE_FACTORS - U77_C1 Genotype:U77 RAW_FILE_NAME=U77_C1.raw SUBJECT_SAMPLE_FACTORS - U77_C2 Genotype:U77 RAW_FILE_NAME=U77_C2.raw SUBJECT_SAMPLE_FACTORS - U44_A1 Genotype:U44 RAW_FILE_NAME=U44_A1.raw SUBJECT_SAMPLE_FACTORS - U44_A2 Genotype:U44 RAW_FILE_NAME=U44_A2.raw SUBJECT_SAMPLE_FACTORS - U44_B1 Genotype:U44 RAW_FILE_NAME=U44_B1.raw SUBJECT_SAMPLE_FACTORS - U44_B2 Genotype:U44 RAW_FILE_NAME=U44_B2.raw SUBJECT_SAMPLE_FACTORS - U44_C1 Genotype:U44 RAW_FILE_NAME=U44_C1.raw SUBJECT_SAMPLE_FACTORS - U44_C2 Genotype:U44 RAW_FILE_NAME=U44_C2.raw SUBJECT_SAMPLE_FACTORS - U41_A1 Genotype:U41 RAW_FILE_NAME=U41_A1.raw SUBJECT_SAMPLE_FACTORS - U41_A2 Genotype:U41 RAW_FILE_NAME=U41_A2.raw SUBJECT_SAMPLE_FACTORS - U41_B2 Genotype:U41 RAW_FILE_NAME=U41_B2.raw SUBJECT_SAMPLE_FACTORS - U41_C1 Genotype:U41 RAW_FILE_NAME=U41_C1.raw SUBJECT_SAMPLE_FACTORS - U41_C2 Genotype:U41 RAW_FILE_NAME=U41_C2.raw SUBJECT_SAMPLE_FACTORS - U25_A1 Genotype:U25 RAW_FILE_NAME=U25_A1.raw SUBJECT_SAMPLE_FACTORS - U25_A2 Genotype:U25 RAW_FILE_NAME=U25_A2.raw SUBJECT_SAMPLE_FACTORS - U25_B1 Genotype:U25 RAW_FILE_NAME=U25_B1.raw SUBJECT_SAMPLE_FACTORS - U25_B2 Genotype:U25 RAW_FILE_NAME=U25_B2.raw SUBJECT_SAMPLE_FACTORS - U25_C1 Genotype:U25 RAW_FILE_NAME=U25_C1.raw SUBJECT_SAMPLE_FACTORS - U25_C2 Genotype:U25 RAW_FILE_NAME=U25_C2.raw SUBJECT_SAMPLE_FACTORS - K1_A1 Genotype:K1 RAW_FILE_NAME=K1_A1.raw SUBJECT_SAMPLE_FACTORS - K1_A2 Genotype:K1 RAW_FILE_NAME=K1_A2.raw SUBJECT_SAMPLE_FACTORS - K1_B1 Genotype:K1 RAW_FILE_NAME=K1_B1.raw SUBJECT_SAMPLE_FACTORS - K1_B2 Genotype:K1 RAW_FILE_NAME=K1_B2.raw SUBJECT_SAMPLE_FACTORS - K1_C1 Genotype:K1 RAW_FILE_NAME=K1_C1.raw SUBJECT_SAMPLE_FACTORS - K1_C2 Genotype:K1 RAW_FILE_NAME=K1_C2.raw SUBJECT_SAMPLE_FACTORS - K2_A1 Genotype:K2 RAW_FILE_NAME=K2_A1.raw SUBJECT_SAMPLE_FACTORS - K2_A2 Genotype:K2 RAW_FILE_NAME=K2_A2.raw SUBJECT_SAMPLE_FACTORS - K2_B1 Genotype:K2 RAW_FILE_NAME=K2_B1.raw SUBJECT_SAMPLE_FACTORS - K2_B2 Genotype:K2 RAW_FILE_NAME=K2_B2.raw SUBJECT_SAMPLE_FACTORS - K2_C1 Genotype:K2 RAW_FILE_NAME=K2_C1.raw SUBJECT_SAMPLE_FACTORS - K2_C2 Genotype:K2 RAW_FILE_NAME=K2_C2.raw SUBJECT_SAMPLE_FACTORS - K3_A1 Genotype:K3 RAW_FILE_NAME=K3_A1.raw SUBJECT_SAMPLE_FACTORS - K3_A2 Genotype:K3 RAW_FILE_NAME=K3_A2.raw SUBJECT_SAMPLE_FACTORS - K3_B1 Genotype:K3 RAW_FILE_NAME=K3_B1.raw SUBJECT_SAMPLE_FACTORS - K3_B2 Genotype:K3 RAW_FILE_NAME=K3_B2.raw SUBJECT_SAMPLE_FACTORS - K3_C1 Genotype:K3 RAW_FILE_NAME=K3_C1.raw SUBJECT_SAMPLE_FACTORS - K3_C2 Genotype:K3 RAW_FILE_NAME=K3_C2.raw #COLLECTION CO:COLLECTION_SUMMARY Coral colonies were brought to the surface intact, and ~3 cm nubbins were CO:COLLECTION_SUMMARY clipped from actively growing branch tips. Nubbins were wrapped in aluminum foil CO:COLLECTION_SUMMARY and immediately frozen in liquid nitrogen. Nubbins were then ground down in an CO:COLLECTION_SUMMARY ice chilled mortar pastel in 10 mL of 2:1 Chloroform/Methanol solution. CO:COLLECTION_SUMMARY Supernatant was then transferred into a test tube labeled with sample I.D. and CO:COLLECTION_SUMMARY "Organic" and vortexed for 10 seconds. 2 mL of .9% NaCl was then added to each CO:COLLECTION_SUMMARY tube and vortexed for an additional 10 seconds. Samples were then allowed to CO:COLLECTION_SUMMARY separate for 15 minutes on ice. After the allotted time, the supernatant was CO:COLLECTION_SUMMARY separated and placed in a separate test tube labeled with sample I.D. and CO:COLLECTION_SUMMARY "Aqueous". Both test tubes were then stored in a -80°C freezer until CO:COLLECTION_SUMMARY processing. CO:SAMPLE_TYPE Tissue and skeleton #TREATMENT TR:TREATMENT_SUMMARY No treatment was applied; study was conducted on natural metabolomic variation TR:TREATMENT_SUMMARY among genotypes #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Metabolomic analyses were performed at the Southeast Center for Integrated SP:SAMPLEPREP_SUMMARY Metabolomics (SECIM) at the University of Florida. Dried powder of aqueous phase SP:SAMPLEPREP_SUMMARY samples acquired from methanol/chloroform extraction were dissolved in 50mM SP:SAMPLEPREP_SUMMARY sodium phosphate buffer with 0.5mM D6-deuterated sodium SP:SAMPLEPREP_SUMMARY trimethylsilylpropanesulfonate (DSS-d6). NMR spectra were measured using the SP:SAMPLEPREP_SUMMARY first slice of a NOESY pulse sequence (tnnoesy) using 14.1 T Bruker Avance II SP:SAMPLEPREP_SUMMARY NMR system with a CP TXI CryoProbe. The acquisition parameters used in Lohr et SP:SAMPLEPREP_SUMMARY al. (2019) and Myer et al. (2020) were utilized to acquire proton spectra. All SP:SAMPLEPREP_SUMMARY spectra were processed and the integrated area was extracted using MestReNova SP:SAMPLEPREP_SUMMARY 11.0-17609 (Mestrelab Research S.L.). Before Fourier transformation, baseline SP:SAMPLEPREP_SUMMARY correction and phase correction were applied with a line-broadening factor of SP:SAMPLEPREP_SUMMARY 0.22 Hz and spectra were normalized with respect to a DSS signal at 0.0 ppm. SP:PROCESSING_METHOD Lyophilization and Homogenization SP:PROCESSING_STORAGE_CONDITIONS -80℃ SP:EXTRACTION_METHOD Modified FOLCH extraction SP:EXTRACT_STORAGE -80℃ SP:SAMPLE_RESUSPENSION In 50 mM phosphate buffer (pH 7.2) with 2 mM EDTA, 0.5 mM DSS and 0.2% sodium SP:SAMPLE_RESUSPENSION azide for aqueous phase samples. SP:SAMPLE_SPIKING 0.5 mM of DSS for aqueous phase samples #ANALYSIS AN:ANALYSIS_TYPE NMR AN:LABORATORY_NAME Matt AN:OPERATOR_NAME Ram Khattri AN:DETECTOR_TYPE Bruker AN:SOFTWARE_VERSION Bruker Topspin AN:ACQUISITION_DATE 02/02/2018 AN:DATA_FORMAT fid, 1r #NMR NM:INSTRUMENT_NAME Bruker Avance Neo 600 MHz/54mm console NM:INSTRUMENT_TYPE FT-NMR NM:NMR_EXPERIMENT_TYPE 1D-1H NM:FIELD_FREQUENCY_LOCK Deuterium NM:STANDARD_CONCENTRATION 0.5 mM DSS NM:SPECTROMETER_FREQUENCY 600.2328273 MHz NM:NMR_PROBE CP TXI CryoProbe NM:NMR_SOLVENT Phosphate buffer (pH 7.2) + 2 mM EDTA + 0.5 mM DSS + 0.2% of sodium azide in NM:NMR_SOLVENT deuterated environment NM:NMR_TUBE_SIZE 1.5 mm O.D. NM:SHIMMING_METHOD Topshim NM:PULSE_SEQUENCE noesypr1d NM:WATER_SUPPRESSION presat NM:PULSE_WIDTH 90-degree NM:RECEIVER_GAIN 256 NM:OFFSET_FREQUENCY 2827.31 Hz NM:CHEMICAL_SHIFT_REF_CPD DSS (4,4-dimethyl-4-silapentane-1-sulfonic acid) NM:TEMPERATURE 25 o C NM:NUMBER_OF_SCANS 64 NM:DUMMY_SCANS 4 NM:ACQUISITION_TIME 4 s NM:RELAXATION_DELAY 1 s NM:SPECTRAL_WIDTH 7211.54 NM:NUM_DATA_POINTS_ACQUIRED 57690 NM:REAL_DATA_POINTS 65536 NM:LINE_BROADENING 0.22 Hz NM:ZERO_FILLING 65,536 points NM:APODIZATION Exponential NM:BASELINE_CORRECTION_METHOD Spline NM:CHEMICAL_SHIFT_REF_STD 0 ppm for DSS NM:BINNED_INCREMENT 0.4ppm NM:BINNED_DATA_EXCLUDED_RANGE greater than 9.5 ppm and below 0.5 ppm including water regions NM:NMR_RESULTS_FILE ST001645_AN002691_Results.txt UNITS:ppm #END