Transcriptomic Response of Juvenile Red King Crab, Paralithodes Camtschaticus, to the Interactive Effects of Ocean Acidification and Warming (NCEI Accession 0170754)

Impacts of elevated carbon dioxide on marine ecosystems depend on physiological responses to consequential decreased pH and increased temperature. Responses to these environmental factors vary among species and life stages, and interactive effects can be significant. To study effects of decreased pH and increased temperature on juvenile red king crab (RKC, Paralithodes camtschaticus) we exposed individuals to three levels of temperature: 11 degrees Celsius (ambient), 13 degrees Celsius, and 14 degrees Celsius, crossed with three levels of pH: 8.0, 7.8 and 7.5, for a total of nine treatments. To better understand the effect of these environmental changes at the level of genome regulation, we analyzed total RNA of whole crabs using Illumina-based RNA-seq whole-transcriptome sequencing. We assembled a RKC transcriptome using Trinity, annotated the transcriptome using Trinotate, and estimated expression levels using bowtie2, samtools and eXpress. Differentially expressed genes were identified using EdgeR. Genes were clustered by expression patterns. Interactive effects were determined by comparing sets of differentially expressed genes using three statistical models to examine the effect of temperature, the effect of pH, and the interaction between temperature and pH in EdgeR. The largest set of differentially expressed genes encoded proteins involved in regulation of extracellular and cuticular structures, including chitin-binding and calcification related proteins.

大气二氧化碳浓度升高对海洋生态系统的影响，取决于生物对随之而来的pH降低与温度升高的生理响应。各类物种乃至同一物种的不同生命阶段，对这些环境因子的响应存在显著差异，且各因子间的交互效应可能极为显著。为探究pH降低与温度升高对幼年红王蟹（red king crab, RKC, *Paralithodes camtschaticus*）的影响，我们将实验个体置于3个温度梯度处理组：11摄氏度（环境对照）、13摄氏度、14摄氏度，并交叉设置3个pH梯度：8.0、7.8与7.5，共计9种实验处理组合。为进一步从基因组调控层面解析此类环境变化的生物学效应，我们采用基于Illumina平台的RNA-seq全转录组测序技术，对整只幼蟹的总RNA进行了测序分析。我们通过Trinity软件组装红王蟹的转录组，利用Trinotate完成转录组功能注释，并借助bowtie2、samtools与eXpress估算基因表达水平。使用EdgeR软件鉴定差异表达基因，并基于基因表达模式对其进行聚类分析。通过EdgeR内置的三种统计模型分别检验温度效应、pH效应以及温度与pH的交互效应，进而通过比较差异表达基因集明确交互效应。差异表达基因中占比最高的一类，编码参与细胞外结构与表皮结构调控的蛋白质，包括几丁质结合蛋白及钙化相关蛋白。