Effects of elevated seawater pCO2 on gene expression patterns in the gills of the green crab, Carcinus maenas

In order to promote our understanding of the responses of green crab acid-base regulatory epithelia to high pCO2, Baltic Sea green crabs were exposed to a pCO2 of 400 Pa for 3 and 7 days after which posterior gills 7 and 9 were sampled. Gills were then subsequently screened for differentially expressed gene transcripts using a 4,462-feature microarray developed by Towle et al. 2010. For each experimental block (gill7-day3, gill7-day7, gill9-day3, gill9-day7), 6 replicate samples were obtained for control (= 39 Pa) and elevated (= 400 Pa) pCO2 exposed animals. Each microarray slide included 4 technical replicates for each transcript and was hybridized with one control pCO2 (labelled with AlexaFluor555) and one elevated pCO2 cDNA (labelled with AlexaFluor647). Lowess-normalized gene expression was calculated as the log2 of the ratio of the fluorescence intensity of the CO2-treatment cDNA to the fluorescence intensity of the control cDNA (log2 ratio=F635/F532).

为加深我们对绿蟹酸碱调节上皮对高二氧化碳分压（pCO₂）响应的认知，本研究将波罗的海绿蟹暴露于400 Pa的pCO₂环境中，分别处理3天与7天，随后采集其第7、9对后鳃组织。后续采用Towle等人2010年开发的包含4462个探针的微阵列（microarray）芯片，筛选差异表达基因转录本。针对每个实验分组（gill7-day3、gill7-day7、gill9-day3、gill9-day7，分别对应第7鳃3天组、第7鳃7天组、第9鳃3天组、第9鳃7天组），本研究为pCO₂对照组（39 Pa）与高pCO₂处理组（400 Pa）各获取6个生物学重复样本。每张微阵列芯片为每个转录本设置4个技术重复，并分别以AlexaFluor555标记对照组pCO₂的cDNA、AlexaFluor647标记处理组pCO₂的cDNA进行杂交反应。采用Lowess归一化法计算基因表达水平，以处理组cDNA荧光强度与对照组cDNA荧光强度的比值的log₂值作为归一化结果（log₂ ratio=F635/F532）。