GSM8187251: GD, pre-EX, subject 2; Homo sapiens; RNA-Seq
Peripheralbloodmononuclearcells(PBMCs)wereisolatedfromwholebloodcollectedbefore-exercise,duringexercise,beforeisoproterenolinfusion,andduringisoproterenolinfusionbydensitygradientcentrifugation(CytivaFicoll-PaqueTM,FisherScientific,Hampton,NH).Gamma-deltaT-cellsfromtherespectiveconditionswereexvivoexpandedfor14dayswithzoledronateandIL-2andfrozenforfutureanalysis.ExpandedV9V2+T-cellswerethawedandresuspendedinPBSandRNAlatersolutionandprocessedbytheUniversityofArizonaGeneticsCoreforbulkRNAsequsingtheIlluminaNextSeq500platform.RNAsamplesqualitywasassessedwithahighsensitivityRNAFragmentAnalyzerKit(advancedAnalytics)andquantitywithanRNAHSassaykit(Qubit).QualitysampleswereusedforlibrarybuildswiththeRapidRNALibraryKit(Swift)andDualCombinatorialIndexingKit(Swift).SampleshadqualityandaveragefragmentsizeassessedwiththeHighSensitivityNGSAnalysisKit(AdvancedAnalytics).QuantitywasassessedwiththeKapaLibraryQuantificationkit(Illumina),andthensampleswereequimolar-pooledandclusteredforsequencingusingIlluminaNextSeq500runchemistry(NextSeq500/550HighOutputv2kit150cycles). ExpandedV9V2+T-cellswerethawedandresuspendedinPBSandRNAlatersolutionandprocessedbytheUniversityofArizonaGeneticsCoreforbulkRNAsequsingtheIlluminaNextSeq500platform.RNAsamplesqualitywasassessedwithahighsensitivityRNAFragmentAnalyzerKit(advancedAnalytics)andquantitywithanRNAHSassaykit(Qubit).QualitysampleswereusedforlibrarybuildswiththeRapidRNALibraryKit(Swift)andDualCombinatorialIndexingKit(Swift).SampleshadqualityandaveragefragmentsizeassessedwiththeHighSensitivityNGSAnalysisKit(AdvancedAnalytics).QuantitywasassessedwiththeKapaLibraryQuantificationkit(Illumina),andthensampleswereequimolar-pooledandclusteredforsequencingusingIlluminaNextSeq500runchemistry(NextSeq500/550HighOutputv2kit150cycles).