Reference genes for transcript quantification in Gracilaria tenuistipitata under drought stress

摘要: Gracilaria tenuistipitata is a red macroalga often found in intertidal environments, where it frequently under drought stress due to tidal water scarcity. In order be able study the molecular mechanisms involved G. resistance dehydration, we have done dehydration assay, tested different RNA extraction protocols, and selected new reference genes for reverse transcription quantitative PCR (RT-qPCR). After testing four methods, that an adapted TRIzol method resulted highest quantities of pure from less biomass. This yielded average 783.6 ng mg−1 clean 25 mg fresh tissue only 1.5 h. Additionally, DNAse I treatment silica spin column purification were prior RT-qPCR. Of nine putative expression stability control drought-like conditions, combination genes: β-GALACTOSIDASE B (GLB), TRANSCRIPTION INITIATION FACTOR IIB (GTF2B), EUKARYOTIC ELONGATION 2 (EEF2), β-TUBULIN (TUB) was selected. allowed us measure level THIOREDOXIN (TRX), OXYGEN-EVOLVING ENHANCER 1 (OEE), NITRATE REDUCTASE (NIA2). TRX upregulated after 1-h kept higher transcriptional levels even 4 h later. contrast, OEE1 slowly downregulated, NIA2 downregulated 1 h but returned regular after. suggests 40 % induced responses without changing relative growth rate or affecting thalli mortality.