#METABOLOMICS WORKBENCH shijuan_20200313_023939 DATATRACK_ID:1942 STUDY_ID:ST001331 ANALYSIS_ID:AN002220 PROJECT_ID:PR000910
VERSION             	1
CREATED_ON             	March 24, 2020, 2:05 pm
#PROJECT
PR:PROJECT_TITLE                 	Rice panicle blast resistence
PR:PROJECT_SUMMARY               	Lipidomics studies of OsGF14b-mediated innate immunity against panicle blast in
PR:PROJECT_SUMMARY               	rice
PR:INSTITUTE                     	Guangdong Academy of Agricultural Sciences
PR:DEPARTMENT                    	Agro-biological Gene Research Center
PR:LAST_NAME                     	Yan
PR:FIRST_NAME                    	Shijuan
PR:ADDRESS                       	No. 20 Jinying Road, Tianhe District, Guangzhou City, Guangdong Province,
PR:ADDRESS                       	510640, China.
PR:EMAIL                         	shijuan@agrogene.ac.cn
PR:PHONE                         	+86-020-38213643
#STUDY
ST:STUDY_TITLE                   	Multi-omics of OsGF14b-mediated innate immunity against panicle blast in rice
ST:STUDY_TITLE                   	(part-II)
ST:STUDY_SUMMARY                 	In the present study, we used a multi-omics approach to decipher the molecular
ST:STUDY_SUMMARY                 	mechanisms of OsGF14b in governing panicle resistance to Magnaporthe
ST:STUDY_SUMMARY                 	oryzae.Results revealed OsGF14b mediated panicle blast resistance was involved
ST:STUDY_SUMMARY                 	in the activation of auxin and JA signaling pathways, resulting in reprogramming
ST:STUDY_SUMMARY                 	of the phenylpropanoid and diterpenoid pathway.
ST:INSTITUTE                     	Agro-biological Gene Research Center , Guangdong Academy of Agricultural
ST:INSTITUTE                     	Sciences
ST:LAST_NAME                     	Yan
ST:FIRST_NAME                    	Shijuan
ST:ADDRESS                       	No. 20 Jinying Road, Tianhe District, Guangzhou City, Guangdong Province,
ST:ADDRESS                       	510640, China.
ST:EMAIL                         	shijuan@agrogene.ac.cn
ST:PHONE                         	+86-020-38213643
#SUBJECT
SU:SUBJECT_TYPE                  	Plant
SU:SUBJECT_SPECIES               	Oryza sativa Japonica Group
SU:TAXONOMY_ID                   	39947
#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           	-	OXGF14b-2-0h-1	Genotype:OsGF14b-overexpression line 2 | Treatment:0h	RAW_FILE_NAME=GF14b-2-0h-1
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-0h-2	Genotype:OsGF14b-overexpression line 2 | Treatment:0h	RAW_FILE_NAME=GF14b-2-0h-2
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-0h-3	Genotype:OsGF14b-overexpression line 2 | Treatment:0h	RAW_FILE_NAME=GF14b-2-0h-3
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-0h-4	Genotype:OsGF14b-overexpression line 2 | Treatment:0h	RAW_FILE_NAME=GF14b-2-0h-4
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-0h-5	Genotype:OsGF14b-overexpression line 2 | Treatment:0h	RAW_FILE_NAME=GF14b-2-0h-5
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-0h-1	Genotype:OsGF14b-overexpression line 4 | Treatment:0h	RAW_FILE_NAME=GF14b-4-0h-1
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-0h-2	Genotype:OsGF14b-overexpression line 4 | Treatment:0h	RAW_FILE_NAME=GF14b-4-0h-2
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-0h-3	Genotype:OsGF14b-overexpression line 4 | Treatment:0h	RAW_FILE_NAME=GF14b-4-0h-3
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-0h-4	Genotype:OsGF14b-overexpression line 4 | Treatment:0h	RAW_FILE_NAME=GF14b-4-0h-4
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-0h-5	Genotype:OsGF14b-overexpression line 4 | Treatment:0h	RAW_FILE_NAME=GF14b-4-0h-5
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-0h-1	Genotype:OsGF14b-overexpression line 6 | Treatment:0h	RAW_FILE_NAME=GF14b-6-0h-1
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-0h-2	Genotype:OsGF14b-overexpression line 6 | Treatment:0h	RAW_FILE_NAME=GF14b-6-0h-2
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-0h-3	Genotype:OsGF14b-overexpression line 6 | Treatment:0h	RAW_FILE_NAME=GF14b-6-0h-3
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-0h-4	Genotype:OsGF14b-overexpression line 6 | Treatment:0h	RAW_FILE_NAME=GF14b-6-0h-4
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-0h-5	Genotype:OsGF14b-overexpression line 6 | Treatment:0h	RAW_FILE_NAME=GF14b-6-0h-5
SUBJECT_SAMPLE_FACTORS           	-	Nip-0h-1	Genotype:Wild-type | Treatment:0h	RAW_FILE_NAME=Nip-0h-1
SUBJECT_SAMPLE_FACTORS           	-	Nip-0h-2	Genotype:Wild-type | Treatment:0h	RAW_FILE_NAME=Nip-0h-2
SUBJECT_SAMPLE_FACTORS           	-	Nip-0h-3	Genotype:Wild-type | Treatment:0h	RAW_FILE_NAME=Nip-0h-3
SUBJECT_SAMPLE_FACTORS           	-	Nip-0h-4	Genotype:Wild-type | Treatment:0h	RAW_FILE_NAME=Nip-0h-4
SUBJECT_SAMPLE_FACTORS           	-	Nip-0h-5	Genotype:Wild-type | Treatment:0h	RAW_FILE_NAME=Nip-0h-5
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-24h-1	Genotype:OsGF14b-overexpression line 2 | Treatment:24h	RAW_FILE_NAME=GF14b-2-24h-1
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-24h-2	Genotype:OsGF14b-overexpression line 2 | Treatment:24h	RAW_FILE_NAME=GF14b-2-24h-2
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-24h-3	Genotype:OsGF14b-overexpression line 2 | Treatment:24h	RAW_FILE_NAME=GF14b-2-24h-3
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-24h-4	Genotype:OsGF14b-overexpression line 2 | Treatment:24h	RAW_FILE_NAME=GF14b-2-24h-4
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-2-24h-5	Genotype:OsGF14b-overexpression line 2 | Treatment:24h	RAW_FILE_NAME=GF14b-2-24h-5
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-24h-1	Genotype:OsGF14b-overexpression line 4 | Treatment:24h	RAW_FILE_NAME=GF14b-4-24h-1
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-24h-5	Genotype:OsGF14b-overexpression line 4 | Treatment:24h	RAW_FILE_NAME=GF14b-4-24h-5
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-24h-2	Genotype:OsGF14b-overexpression line 4 | Treatment:24h	RAW_FILE_NAME=GF14b-4-24h-2
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-24h-3	Genotype:OsGF14b-overexpression line 4 | Treatment:24h	RAW_FILE_NAME=GF14b-4-24h-3
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-4-24h-4	Genotype:OsGF14b-overexpression line 4 | Treatment:24h	RAW_FILE_NAME=GF14b-4-24h-4
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-24h-1	Genotype:OsGF14b-overexpression line 6 | Treatment:24h	RAW_FILE_NAME=GF14b-6-24h-1
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-24h-2	Genotype:OsGF14b-overexpression line 6 | Treatment:24h	RAW_FILE_NAME=GF14b-6-24h-2
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-24h-3	Genotype:OsGF14b-overexpression line 6 | Treatment:24h	RAW_FILE_NAME=GF14b-6-24h-3
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-24h-4	Genotype:OsGF14b-overexpression line 6 | Treatment:24h	RAW_FILE_NAME=GF14b-6-24h-4
SUBJECT_SAMPLE_FACTORS           	-	OXGF14b-6-24h-5	Genotype:OsGF14b-overexpression line 6 | Treatment:24h	RAW_FILE_NAME=GF14b-6-24h-5
SUBJECT_SAMPLE_FACTORS           	-	Nip-24h-1	Genotype:Wild-type | Treatment:24h	RAW_FILE_NAME=Nip-24h-1
SUBJECT_SAMPLE_FACTORS           	-	Nip-24h-2	Genotype:Wild-type | Treatment:24h	RAW_FILE_NAME=Nip-24h-2
SUBJECT_SAMPLE_FACTORS           	-	Nip-24h-3	Genotype:Wild-type | Treatment:24h	RAW_FILE_NAME=Nip-24h-3
SUBJECT_SAMPLE_FACTORS           	-	Nip-24h-4	Genotype:Wild-type | Treatment:24h	RAW_FILE_NAME=Nip-24h-4
SUBJECT_SAMPLE_FACTORS           	-	Nip-24h-5	Genotype:Wild-type | Treatment:24h	RAW_FILE_NAME=Nip-24h-5
#COLLECTION
CO:COLLECTION_SUMMARY            	The panicles at the initial heading stage of the wild-type Nipponbare (Nip) and
CO:COLLECTION_SUMMARY            	OsGF14b-overexpressing plants were harvested before (Nip-0h; OXGF14b-2-0h;
CO:COLLECTION_SUMMARY            	OXGF14b-4-0h; OXGF14b-6-0h) and after M. oryzae 24-hour inoculation (Nip-24h;
CO:COLLECTION_SUMMARY            	OXGF14b-2-24h; OXGF14b-4-24h; OXGF14b-6-24h) respectively. They were immediately
CO:COLLECTION_SUMMARY            	frozen in liquid nitrogen, with each biological replicate containing panicle
CO:COLLECTION_SUMMARY            	pooled from 10 individual plants.
CO:SAMPLE_TYPE                   	Seeds
#TREATMENT
TR:TREATMENT_SUMMARY             	Wild-type japonica rice (Oryzae sativa cv. Nipponbare) and three OsGF14b gene
TR:TREATMENT_SUMMARY             	overexpressing lines, including transgenic line 2 (OXGF14b-2), transgenic line 4
TR:TREATMENT_SUMMARY             	(OXGF14b-4), transgenic line 6 (OXGF14b-6) were used in this study. Rice seeds
TR:TREATMENT_SUMMARY             	were surface-sterilized and transferred to 1/2 MS medium and incubated in a
TR:TREATMENT_SUMMARY             	growth chamber for germination under light of 200 μmol/m2/s with a 12-h
TR:TREATMENT_SUMMARY             	photoperiod at 26℃. Subsequently, rice seedlings were transplanted into soil
TR:TREATMENT_SUMMARY             	and kept in a greenhouse. M. oryzae GD08-T13 was used for rice blast
TR:TREATMENT_SUMMARY             	inoculation.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	The rice panicle pre-cooled in liquid nitrogen were ground using a Mixer/mill
SP:SAMPLEPREP_SUMMARY            	(MM400; Retsch) with steel ball for 30 seconds at 30 HZ. Fifty milligram of rice
SP:SAMPLEPREP_SUMMARY            	panicle powder of each sample was extracted with a fixed volume (1 ml) of
SP:SAMPLEPREP_SUMMARY            	pre-cooled (−20 °C) extraction solvent (methanol: chloroform: water = 5: 2:
SP:SAMPLEPREP_SUMMARY            	2) was added to homogenized tissues. After adding the extraction solvent, the
SP:SAMPLEPREP_SUMMARY            	vials/tubes were thoroughly vortexed for 1 min and then incubated on an orbital
SP:SAMPLEPREP_SUMMARY            	shaker (200 rpm) for 10 min at 4 °C followed by a 15 min sonication step.
SP:SAMPLEPREP_SUMMARY            	For phase separation, a volume of 500 µl of solvent (methanol: water = 1: 3),
SP:SAMPLEPREP_SUMMARY            	was added to each vial/tube and the samples were again thoroughly vortexed for
SP:SAMPLEPREP_SUMMARY            	1 min. After that, the samples are centrifuged at a speed of 14000rpm for
SP:SAMPLEPREP_SUMMARY            	10 min at 4 °C. one fixed volume 500 μL of the unpolar phase (the upper
SP:SAMPLEPREP_SUMMARY            	phase) was transferred into pre-labeled 1.5 ml microcentrifuge tube. Then the
SP:SAMPLEPREP_SUMMARY            	samples were dried in a SpeedVac concentrator without heating. The dried 500 μL
SP:SAMPLEPREP_SUMMARY            	aliquot from the upper phase for lipids profiling was re-suspended in 200 μL
SP:SAMPLEPREP_SUMMARY            	UPLC-grade ACN: 2-propanol: dichloromethane (1:1:1, v/v/v) and analyzed using
SP:SAMPLEPREP_SUMMARY            	LC-MS for lipidomics study.
SP:PROCESSING_STORAGE_CONDITIONS 	-80℃
SP:EXTRACT_STORAGE               	-80℃
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_SUMMARY        	Five μL of each sample was eluted using a CORTECS® C18 column (150 mm × 2.1
CH:CHROMATOGRAPHY_SUMMARY        	mm, 2.7μm, Waters) with 0.4 mL/min flow rate. The mobile phase A was water: ACN
CH:CHROMATOGRAPHY_SUMMARY        	(40: 60, v/v) with 0.1% formic acid and 10mM ammonium formate, and the mobile
CH:CHROMATOGRAPHY_SUMMARY        	phase B was 2-propanol: ACN (90:10, v/v) with 0.1% formic acid and 10mM ammonium
CH:CHROMATOGRAPHY_SUMMARY        	formate. The compounds were separated by a elution gradient: 20% B was firstly
CH:CHROMATOGRAPHY_SUMMARY        	maintained for 0.2 min, then linearly decreased to 60% B from 0.2 to 2min, to
CH:CHROMATOGRAPHY_SUMMARY        	100% B from 2 to 9min, and maintained at 100% B from 9 to 10 min, then linearly
CH:CHROMATOGRAPHY_SUMMARY        	increased to 20% B from 10 to 10.5 min followed by equilibration at 20% B for
CH:CHROMATOGRAPHY_SUMMARY        	3.5 min.
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Thermo Dionex Ultimate 3000
CH:COLUMN_NAME                   	Waters CORTECS C18 (150 mm×2.1 mm, 2.7µm)
CH:FLOW_RATE                     	0.4 mL/min
CH:SOLVENT_A                     	Water: Acetonitrile(40: 60, v/v) with0.1% formic acid and 10mM ammonium formate
CH:SOLVENT_B                     	2-propanol: Acetonitrile (90:10, v/v) with 0.1% formic acid and 10mM ammonium
CH:SOLVENT_B                     	formate
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Thermo Fusion Tribrid Orbitrap
MS:INSTRUMENT_TYPE               	Orbitrap
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	NEGATIVE
MS:MS_COMMENTS                   	The spray voltage was set to -3000 V in the negative-ion modes, with the
MS:MS_COMMENTS                   	following ion-source properties: sheath gas, 45 Arbs; auxiliary gas, 10 Arbs;
MS:MS_COMMENTS                   	sweep gas, 0 Arbs; ion-transfer tube temperature, 320 °C; vaporizer temp, 350
MS:MS_COMMENTS                   	°C. All FTMS data were acquired using the following conditions: detector type,
MS:MS_COMMENTS                   	orbitrap; orbitrap resolution, 120000; scan range was set to m/z 200-1200. The
MS:MS_COMMENTS                   	AGC was set at 5.0 e5 and the maximum injection time was set to 100 ms. RF lens
MS:MS_COMMENTS                   	was set to 60%, and the microscans was 1. data type, profile. All FTMS2 data
MS:MS_COMMENTS                   	were acquired using the following conditions: isolation mode, quadrupole;
MS:MS_COMMENTS                   	isolation window, 1 m/z; detector type, orbitrap; scan range, auto; AGC target,
MS:MS_COMMENTS                   	5.0 e4; maximum injection time, 100 ms; microscans, 1; orbitrap resolution,
MS:MS_COMMENTS                   	30000; first mass, 50 m/z; data type, profile. The HCD collision energy was set
MS:MS_COMMENTS                   	to 25% with ± HCD collision energy was set 5% in positive mode, and the HCD
MS:MS_COMMENTS                   	collision energy was set to 30% with ± HCD collision energy was set 5% in
MS:MS_COMMENTS                   	negative mode. All data was acquired by the Xcalibur 4.1 software (Thermo Fisher
MS:MS_COMMENTS                   	Scientific, USA). Thermo Scientific™ LipidSearch™ 5.0 software was used for
MS:MS_COMMENTS                   	lipidome data processing.
MS:MS_RESULTS_FILE               	ST001331_AN002220_Results.txt	UNITS:Relative content	Has m/z:Yes	Has RT:Yes	RT units:Minutes
#END