#METABOLOMICS WORKBENCH pramodwangikar_20190507_010552 DATATRACK_ID:1721 STUDY_ID:ST001180 ANALYSIS_ID:AN001958 PROJECT_ID:PR000792 VERSION 1 CREATED_ON May 8, 2019, 9:50 am #PROJECT PR:PROJECT_TITLE Metabolome Analysis of Synechococcus elongatus PCC 11802 PR:PROJECT_TYPE Metabolomics PR:PROJECT_SUMMARY Metabolomics Analysis of a novel freshwater cyanobacterium, Synechococcus PR:PROJECT_SUMMARY elongatus PCC 11802 isolated by us from Powai Lake, Mumbai, India. PCC 11802 PR:PROJECT_SUMMARY cells were grown under ambient and 1% CO2 conditions and metabolomics data was PR:PROJECT_SUMMARY collected in three biological replicates and two technical replicates (n=6). The PR:PROJECT_SUMMARY study aims to find metabolomics changes in this cyanobacterium at elevated CO2 PR:PROJECT_SUMMARY levels. PR:INSTITUTE Indian Institute of Technology-Bombay PR:DEPARTMENT Department of Chemical Engineering PR:LABORATORY Biosystems Engineering Lab PR:LAST_NAME Wangikar PR:FIRST_NAME Pramod PR:ADDRESS Room No. 136, Biosystems Engineering Laboratory, Mumbai, Maharashtra, 400076, PR:ADDRESS India PR:EMAIL wangikar@iitb.ac.in PR:PHONE 2225764215 PR:FUNDING_SOURCE DBT-PAN IIT Center for Bioenergy (Grant No: BT/EB/PAN IIT/2012) #STUDY ST:STUDY_TITLE Metabolome Profiling of Synechococcus elogatus PCC 11802 ST:STUDY_TYPE Quantitative Metabolomics ST:STUDY_SUMMARY Metabolomics Analysis of a novel freshwater cyanobacterium, Synechococcus ST:STUDY_SUMMARY elongatus PCC 11802 isolated by us from Powai Lake, Mumbai, India. PCC 11802 ST:STUDY_SUMMARY cells were grown under ambient and 1% CO2 conditions and metabolomics data was ST:STUDY_SUMMARY collected in three biological replicates and two technical replicates (n=6). The ST:STUDY_SUMMARY study aims to find metabolomics changes in this cyanobacterium at elevated CO2 ST:STUDY_SUMMARY levels. ST:INSTITUTE Indian Institute of Technology Bombay ST:DEPARTMENT Department of Chemical Engineering ST:LABORATORY Biosystems Engineering Lab ST:LAST_NAME Wangikar ST:FIRST_NAME Pramod P ST:ADDRESS Powai ST:EMAIL wangikar@iitb.ac.in ST:PHONE +91 (22) 2576 7232 #SUBJECT SU:SUBJECT_TYPE Bacteria SU:SUBJECT_SPECIES Synechococcus elongatus PCC 11802 SU:TAXONOMY_ID 1131 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - PCC 11802 AMBIENT CO2 R1 Conditions:Ambient CO2 SUBJECT_SAMPLE_FACTORS - PCC 11802 AMBIENT CO2 R2 Conditions:Ambient CO2 SUBJECT_SAMPLE_FACTORS - PCC 11802 AMBIENT CO2 R3 Conditions:Ambient CO2 SUBJECT_SAMPLE_FACTORS - PCC 11802 1 PERCENT CO2 R1 Conditions:1% CO2 SUBJECT_SAMPLE_FACTORS - PCC 11802 1 PERCENT CO2 R2 Conditions:1% CO2 SUBJECT_SAMPLE_FACTORS - PCC 11802 1 PERCENT CO2 R3 Conditions:1% CO2 #COLLECTION CO:COLLECTION_SUMMARY The PCC 11802 was grown under ambient and 1% CO2 conditions in triplicates in CO:COLLECTION_SUMMARY shake flasks at 200 uE light intensity, 38 degrees C temperature, 120 rpm. The CO:COLLECTION_SUMMARY samples were collected in exponential phase (OD730 = 0.6).The metabolites were CO:COLLECTION_SUMMARY extracted using methanol-chloroform-water method as described in "Metabolite CO:COLLECTION_SUMMARY Extraction Protocol" file. The samples were stored in a -80 degrees C deep CO:COLLECTION_SUMMARY freezer until analyzed using LCMS. CO:SAMPLE_TYPE Cultured cells #TREATMENT TR:TREATMENT_SUMMARY The metabolites were extracted using a methanol-chloroform-water method TR:TREATMENT_SUMMARY described in the "Metabolite Extraction" file of the collection data. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The metabolite extracts of different collected samples were reconstituted in 100 SP:SAMPLEPREP_SUMMARY μL of 50/50 methanol-water mixture. An equal amount of fully 13C-labeled SP:SAMPLEPREP_SUMMARY biomass extract from cyanobacterium Synechococcus elongatus PCC 11801 was added SP:SAMPLEPREP_SUMMARY as internal standard. The mixture is vortexed. The samples are filtered through SP:SAMPLEPREP_SUMMARY syringe filters. Injection volume was 6 uL. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Shimadzu 20AD CH:COLUMN_NAME Phenomenex Synergi Hydro RP 100 A (100 x 2mm, 2.5um) CH:FLOW_GRADIENT 0% B (0 min), 0% B (2min), 35% B (8 min), 35% B (10.5 min), 90% B (15.50 min), CH:FLOW_GRADIENT 90% B (20.5 min) and 0% B (22 min) CH:FLOW_RATE 0.3 mL/minute CH:SOLVENT_A 10 mM tributylamine + 15mM acetic acid in water CH:SOLVENT_B 100% Methanol #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME Biosystems Engineering Lab AN:OPERATOR_NAME Damini Jaiswal AN:SOFTWARE_VERSION Analyst #MS MS:INSTRUMENT_NAME ABI Sciex 5600+ TripleTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE NEGATIVE MS:MS_COMMENTS Data Acquisition was done using IDA method with MS/MS trigerred for top 10 MS:MS_COMMENTS abundant precursor ions. Cycle time = 1s MS:COLLISION_ENERGY -30 MS:ION_SOURCE_TEMPERATURE 450 #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS Area Ratio MS_METABOLITE_DATA_START Samples PCC 11802 AMBIENT CO2 R1 PCC 11802 AMBIENT CO2 R1 PCC 11802 AMBIENT CO2 R2 PCC 11802 AMBIENT CO2 R2 PCC 11802 AMBIENT CO2 R3 PCC 11802 AMBIENT CO2 R3 PCC 11802 1 PERCENT CO2 R1 PCC 11802 1 PERCENT CO2 R1 PCC 11802 1 PERCENT CO2 R2 PCC 11802 1 PERCENT CO2 R2 PCC 11802 1 PERCENT CO2 R3 PCC 11802 1 PERCENT CO2 R3 Factors Conditions:Ambient CO2 Conditions:Ambient CO2 Conditions:Ambient CO2 Conditions:Ambient CO2 Conditions:Ambient CO2 Conditions:Ambient CO2 Conditions:1% CO2 Conditions:1% CO2 Conditions:1% CO2 Conditions:1% CO2 Conditions:1% CO2 Conditions:1% CO2 3-phosphoglyceric acid 0.8226 0.8134 0.9449 0.9358 0.8654 0.9137 1.695 1.702 0.863 0.8486 1.35 1.387 Fructose 1,6 bisphosphate 0.793 0.7992 0.7864 0.7365 0.8006 0.7609 2.345 2.438 1.71 1.476 1.885 2.105 Fructose-6-phosphate 0.4519 0.3863 0.5698 0.6851 0.7438 0.7478 2.253 2.364 1.635 1.626 2.497 2.45 Glucose-6-phosphate 0.4184 0.4322 0.6088 0.5329 0.6352 0.5702 2.031 2.048 1.537 1.33 2.347 2.407 Sedoheptulose 1,7 bisphosphate 0.00367 0.01864 0.01816 0.02794 0.02016 0.006 0.1246 0.1035 0.09343 0.1004 0.1376 0.1534 Sedoheptulose-7-phosphate 0.5494 0.5644 0.7189 0.7299 0.7159 0.7973 2.009 1.982 1.38 1.374 2.301 2.341 Ribulose 1,5 bisphosphate 0.759 0.8297 1.283 1.348 1.856 1.682 1.272 1.249 0.6306 0.5965 1.045 1.07 Glycerol-3-phosphate 2.993 3.783 3.348 4.093 3.123 2.894 2.496 1.706 1.253 1.688 2.65 2.893 ADP-glucose 11.29 16.72 14.91 17.64 17.84 17.47 8.701 11.7 9.58 12.94 17.74 18.9 UDP-glucose 2.371 2.334 2.731 3.762 3.764 3.801 6.06 4.957 4.427 5.088 6.087 6.955 Phosphoenolpyruvic acid 1.137 1.076 1.197 1.138 0.9881 0.8846 1.23 1.195 0.4488 0.5165 0.7523 0.7199 Acetyl Coenzyme A 26.73 31.34 19.68 31.55 30.14 28.39 18.99 23.59 14.39 23.74 26.86 14.08 Aspartic acid 0.4578 0.6391 0.5844 0.7933 0.6114 0.8224 1.784 2.097 0.9002 1.102 1.38 1.64 Glutamic acid 0.8688 0.8876 1.069 1.069 1.108 1.14 1.278 1.283 1.123 1.147 1.642 1.664 Glutamine 0.8386 0.8171 0.9045 0.9241 1.2 1.179 1.279 1.32 0.8459 0.9202 1.187 1.384 Succinic acid 3.192 3.053 2.624 3.417 1.853 3.047 3.457 5.766 2.108 4.379 2.376 3.238 Sucrose-6-phosphate 0.1991 0.2191 0.1948 0.1712 0.285 0.3103 2.869 2.965 4.12 4.113 6.137 6.686 Sucrose 0.2507 0.2779 0.2652 0.2505 0.465 0.5216 1.088 0.9409 1.633 1.553 1.856 1.914 ATP 1.306 1.368 1.305 1.277 1.469 1.453 2.397 2.415 1.408 1.415 2.222 2.197 ADP 1.341 1.373 1.25 1.386 1.431 1.433 2.424 2.408 1.336 1.381 2.108 2.057 AMP 1.071 1.074 1.581 1.634 1.61 1.634 2.921 3.077 2.677 2.696 3.513 3.629 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name quantified m/z KEGG ID 3-phosphoglyceric acid 184.98 C00197 Fructose 1,6 bisphosphate 338.96 C05378 Fructose-6-phosphate 259.02 C00085 Glucose-6-phosphate 259.02 C00668 Sedoheptulose 1,7 bisphosphate 369.01 C00447 Sedoheptulose-7-phosphate 289.03 C05382 Ribulose 1,5 bisphosphate 308.97 C01182 Glycerol-3-phosphate 171 C00093 ADP-glucose 588.07 C00498 UDP-glucose 565.05 C00029 Phosphoenolpyruvic acid 166.97 C00074 Acetyl Coenzyme A 808.12 C00024 Aspartic acid 132.03 C00049 Glutamic acid 146.04 C00025 Glutamine 145.08 C00064 Succinic acid 117.02 C00042 Sucrose-6-phosphate 421.07 C16688 Sucrose 341.11 C00089 ATP 505.98 C00002 ADP 426.02 C00008 AMP 346.05 C00020 METABOLITES_END #END