#METABOLOMICS WORKBENCH Karin_20200911_055403_mwtab.txt DATATRACK_ID:2168 STUDY_ID:ST001486 ANALYSIS_ID:AN002463 PROJECT_ID:000000 VERSION 1 CREATED_ON September 15, 2020, 12:43 pm #PROJECT PR:PROJECT_TITLE Ndufs4 KO mouse model metabolomics studies PR:PROJECT_TYPE Multi-platform metabolomics analysis PR:PROJECT_SUMMARY Multi-platform metabolomics analysis of tissues and biofluids from the Ndufs4 PR:PROJECT_SUMMARY knockout (Ndufs4-/-) mouse model of human Leigh syndrome PR:INSTITUTE North-West University PR:LAST_NAME Louw PR:FIRST_NAME Roan PR:ADDRESS Hofman Street PR:EMAIL Roan.Louw@nwu.ac.za PR:PHONE +27 18 299 4074 #STUDY ST:STUDY_TITLE Metabolomics of Ndufs4 KO brain regions (part - VI) ST:STUDY_SUMMARY Untargeted GC-TOF-MS analysis of Ndufs4 KO and WT mouse cerebellum (CB) ST:INSTITUTE North-West University ST:LAST_NAME Louw ST:FIRST_NAME Roan ST:ADDRESS Hofman Street ST:EMAIL Roan.Louw@nwu.ac.za ST:PHONE +27 18 299 4074 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN Ndufs4, https://www.jax.org/strai n/02705 8 SU:AGE_OR_AGE_RANGE 45-50 days SU:GENDER Male SU:ANIMAL_ANIMAL_SUPPLIER Jackson Laboratory (ME, USA) SU:ANIMAL_LIGHT_CYCLE 12:12 h SU:ANIMAL_FEED Rodent Breeder, Cat. #RM1845, LabChef, Nutritionhub SU:ANIMAL_WATER ad libitum #FACTORS #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 - CBA 1 (25) Genotype:WT RAW_FILE_NAME=CBA 1 (25)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 2 (26) Genotype:WT RAW_FILE_NAME=CBA 2 (26)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 4 (28) Genotype:WT RAW_FILE_NAME=CBA 4 (28)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 6 (30) Genotype:WT RAW_FILE_NAME=CBA 6 (30)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 9 (1) Genotype:WT RAW_FILE_NAME=CBA 9 (1)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 10 (2) Genotype:WT RAW_FILE_NAME=CBA 10 (2)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 12 (4) Genotype:WT RAW_FILE_NAME=CBA 12 (4)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 14 (5) Genotype:WT RAW_FILE_NAME=CBA 14 (5)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 16 (7) Genotype:WT RAW_FILE_NAME=CBA 16 (7)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 17 (8) Genotype:WT RAW_FILE_NAME=CBA 17 (8)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 18 (9) Genotype:WT RAW_FILE_NAME=CBA 18 (9)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 19 (11) Genotype:WT RAW_FILE_NAME=CBA 19 (11)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 22 (37) Genotype:WT RAW_FILE_NAME=CBA 22 (37)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 24 (39) reg Genotype:WT RAW_FILE_NAME=CBB 24 (39) reg_1.peg SUBJECT_SAMPLE_FACTORS - CBB 25 (40) Genotype:WT RAW_FILE_NAME=CBB 25 (40)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 28 (E6) Genotype:WT RAW_FILE_NAME=CBB 28 (E6)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 31 (E3) Genotype:WT RAW_FILE_NAME=CBB 31 (E3)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 34 (12) Genotype:WT RAW_FILE_NAME=CBB 34 (12)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 35 (13) Genotype:WT RAW_FILE_NAME=CBB 35 (13)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 38 (17) Genotype:WT RAW_FILE_NAME=CBB 38 (17)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 39 (18) Genotype:WT RAW_FILE_NAME=CBB 39 (18)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 41 (20) Genotype:WT RAW_FILE_NAME=CBB 41 (20)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 43 (22) Genotype:WT RAW_FILE_NAME=CBB 43 (22)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 44 (23) Genotype:WT RAW_FILE_NAME=CBB 44 (23)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 3 (27) Genotype:KO RAW_FILE_NAME=CBA 3 (27)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 5 (29) Genotype:KO RAW_FILE_NAME=CBA 5 (29)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 7 (32) Genotype:KO RAW_FILE_NAME=CBA 7 (32)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 8 (33) Genotype:KO RAW_FILE_NAME=CBA 8 (33)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 11 (3) Genotype:KO RAW_FILE_NAME=CBA 11 (3)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 13 (34) Genotype:KO RAW_FILE_NAME=CBA 13 (34)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 15 (35) Genotype:KO RAW_FILE_NAME=CBA 15 (35)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 20 (14) Genotype:KO RAW_FILE_NAME=CBA 20 (14)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 21 (36) Genotype:KO RAW_FILE_NAME=CBA 21 (36)_1.peg SUBJECT_SAMPLE_FACTORS - CBA 23 (38) Genotype:KO RAW_FILE_NAME=CBA 23 (38)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 26 (41) Genotype:KO RAW_FILE_NAME=CBB 26 (41)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 27 (42) Genotype:KO RAW_FILE_NAME=CBB 27 (42)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 29 (E1) Genotype:KO RAW_FILE_NAME=CBB 29 (E1)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 30 (E2) Genotype:KO RAW_FILE_NAME=CBB 30 (E2)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 32 (E4) Genotype:KO RAW_FILE_NAME=CBB 32 (E4)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 33 (E5) Genotype:KO RAW_FILE_NAME=CBB 33 (E5)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 36 (15) Genotype:KO RAW_FILE_NAME=CBB 36 (15)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 37 (16) Genotype:KO RAW_FILE_NAME=CBB 37 (16)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 40 (19) Genotype:KO RAW_FILE_NAME=CBB 40 (19)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 42 (21) Genotype:KO RAW_FILE_NAME=CBB 42 (21)_1.peg SUBJECT_SAMPLE_FACTORS - CBB 45 (24) Genotype:KO RAW_FILE_NAME=CBB 45 (24)_1.peg #COLLECTION CO:COLLECTION_SUMMARY Mice were euthanized between postnatal day (P) 45-50 via cervical dislocation at CO:COLLECTION_SUMMARY the same time of day (8:00-9:00 AM) after overnight (12-h) fasting. The brain CO:COLLECTION_SUMMARY was removed and rinsed with saline solution (SABAX PBS; 0.9% NaCl (w/v), #7634, CO:COLLECTION_SUMMARY Adcock Ingram) to remove surrounding blood. The brain regions of interest, CO:COLLECTION_SUMMARY namely the anterior cortex (AC), brainstem (BS), cerebellum (CB) and olfactory CO:COLLECTION_SUMMARY bulbs (OB), were then dissected, snap-frozen in liquid nitrogen (within 15 CO:COLLECTION_SUMMARY minutes postmortem) and stored at − 80°C until used. CO:SAMPLE_TYPE Brain #TREATMENT TR:TREATMENT_SUMMARY The animals did not receive any treatment #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Brain regions were homogenized in the presence of internal standards using a SP:SAMPLEPREP_SUMMARY vibration mill. Metabolite extraction was achieved using a modified monophasic SP:SAMPLEPREP_SUMMARY Bligh–Dyer extraction method with a solvent ratio of 3:1:1 SP:SAMPLEPREP_SUMMARY (methanol:water:chloroform). SP:SAMPLEPREP_PROTOCOL_FILENAME GC_sample_prep_protocol.pdf;Data_processing_method.pdf #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 7890A CH:COLUMN_NAME Restek Rtx-5Sil MS (30 x 0.25mm, 0.25um) #ANALYSIS AN:ANALYSIS_TYPE MS AN:ANALYSIS_PROTOCOL_FILE Data_processing_method.pdf #MS MS:INSTRUMENT_NAME Leco Pegasus HT TOF MS:INSTRUMENT_TYPE GC-TOF MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS The LECO Corporation ChromaTOF® software (v 4.5x) was used for data acquisition MS:MS_COMMENTS and extraction. This included automatic baseline removal via the “spanning” MS:MS_COMMENTS tracking method (offset of 1; just above the noise) and auto smoothing, with the MS:MS_COMMENTS software’s Statistical Compare feature used to align peaks. Spectral matching MS:MS_COMMENTS was done using the NIST11 commercial library and an in-house mass spectral MS:MS_COMMENTS library in order to identify important analytes. MS:MS_RESULTS_FILE ST001486_AN002463_Results.txt UNITS:Area Has m/z:Yes Has RT:Yes RT units:Seconds #END