Identification and selection of optimal reference genes for qPCR-based gene expression analysis in Fucus distichus under various abiotic stresses

Quantitative gene expression analysis is an important tool in the scientist’s belt. The identification of evenly expressed reference genes is necessary for accurate quantitative gene expression analysis, whether by traditional RT-PCR (reverse-transcription polymerase chain reaction) or by qRT-PCR (quantitative real-time PCR; qPCR). In the Stramenopiles (the major line of eukaryotes that includes brown algae) there is a noted lack of known reference genes for such studies, largely due to the absence of available molecular tools.  Here we present a set of nine reference genes (Elongation Factor 1 alpha (EF1A), Elongation Factor 2 alpha (EF2A), Elongation Factor 1 beta (EF1B), 14-3-3 Protein, Ubiquitin Conjugating Enzyme (UBCE2), Glyceraldehyde-3-phosphate Dehydrogenase (GAPDH), Actin Related Protein Complex (ARP2/3), Ribosomal Protein (40s; S23), and Actin) for the brown alga Fucus distichus. These reference genes were tested on adult sporophytes across six abiotic stress conditions (desi...

定量基因表达分析是科研工作者的核心研究工具之一。无论采用传统逆转录聚合酶链式反应（reverse-transcription polymerase chain reaction, RT-PCR）还是实时定量聚合酶链式反应（quantitative real-time PCR, qRT-PCR；qPCR），要实现精准的定量基因表达分析，均需鉴定出表达稳定性一致的内参基因。在包含褐藻在内的真核生物主要类群异鞭毛类（Stramenopiles）中，此类研究的已知内参基因极度匮乏，这主要是由于缺乏可用的分子研究工具。本研究为褐藻墨角藻（Fucus distichus）筛选得到一套共9个内参基因，分别为延伸因子1α（Elongation Factor 1 alpha, EF1A）、延伸因子2α（Elongation Factor 2 alpha, EF2A）、延伸因子1β（Elongation Factor 1 beta, EF1B）、14-3-3蛋白、泛素结合酶（Ubiquitin Conjugating Enzyme, UBCE2）、甘油醛-3-磷酸脱氢酶（Glyceraldehyde-3-phosphate Dehydrogenase, GAPDH）、肌动蛋白相关蛋白复合体（Actin Related Protein Complex, ARP2/3）、核糖体蛋白（40S亚基；S23）以及肌动蛋白。本研究针对成年孢子体在六种非生物胁迫条件下对这些内参基因进行了测试（desi...）