Institute of Evolution and Marine Biodiversity, Ocean University of China, Qingdao, China; Key Laboratory of Mariculture (Ocean University of China), Qingdao, Ministry of Education, China.
Environ Toxicol Pharmacol. 2018 Jun;60:76-81. doi: 10.1016/j.etap.2018.04.010. Epub 2018 Apr 14.
High concentration of ammonia is a common issue in the aquaculture industry and often causes detrimental effects to aquatic products. Exploring expression regulation of genes involved in acute ammonia stress can help to understand the molecular mechanisms of ammonia response. Quantitative real-time PCR (qRT-PCR) with proper reference genes is an effective way to normalize the expression of target genes. To identify the most suitable reference genes for Octopus minor (Mollusca: Cephalopoda: Octopoda) under acute ammonia stress and the normal culturing, nine candidate genes were selected for the validation: OD, RPS18b, RPL29, RPS5, EF-1α, RPL6, AA4, ACT, TUB. The results showed that the stability of candidate genes varied considerably in gill, digestive gland, brain and hemolymph. Thus, the reference genes were determined separately in different tissues. RPS5 and RPL6 showed relatively high stability in gill, while RPL6, TUB, RPL29 and OD were four suitable reference genes in digestive gland. EF-1α, TUB and RPL6 were the best combination in brain and EF-1α and RPS18b were the most appropriate reference genes in hemolymph.
高浓度的氨是水产养殖行业中的一个常见问题，通常会对水产养殖产品造成有害影响。探索参与急性氨胁迫的基因的表达调控可以帮助理解氨反应的分子机制。使用适当的内参基因进行定量实时 PCR(qRT-PCR)是标准化靶基因表达的有效方法。为了鉴定在急性氨胁迫和正常培养下的短蛸（软体动物门：头足纲：八腕目）中最适合的参考基因，选择了 9 个候选基因进行验证：OD、RPS18b、RPL29、RPS5、EF-1α、RPL6、AA4、ACT、TUB。结果表明，候选基因在鳃、消化腺、脑和血淋巴中的稳定性差异很大。因此，在不同组织中分别确定了参考基因。RPS5 和 RPL6 在鳃中表现出相对较高的稳定性，而 RPL6、TUB、RPL29 和 OD 是消化腺中四个合适的参考基因。EF-1α、TUB 和 RPL6 是脑的最佳组合，EF-1α和 RPS18b 是血淋巴中最适合的参考基因。
