Transcriptomic profiling of Aeromonas hydrophila under nanoplastic and shear stress exposure

For transcriptomic analysis, Aeromonas hydrophila cells from each treatment group were collected from three biological replicates. Cells were harvested by centrifugation, washed, and total RNA was extracted following the Majorbio protocol. After DNase I treatment, RNA purity was assessed using NanoDrop 2000. Strand-specific cDNA libraries were prepared using the TruSeq RNA Sample Prep Kit (Illumina). Briefly, mRNA was fragmented, cDNA synthesized using random hexamers, and dUTP was incorporated during second-strand synthesis. After adapter ligation, uracil-N-glycosylase was used to remove the second strand, followed by PCR amplification. Sequencing was conducted on the Illumina NovaSeq 6000 platform with paired-end reads. Differential expression analysis was performed using DESeq2, and KEGG pathway enrichment was conducted via the Majorbio Cloud Platform.

为开展转录组分析，从各处理组中收集3份生物学重复的嗜水气单胞菌（Aeromonas hydrophila）细胞。通过离心收集细胞并洗涤后，遵循美吉生物（Majorbio）的操作规范提取总RNA。经脱氧核糖核酸酶I（DNase I）处理后，使用NanoDrop 2000检测RNA纯度。采用Illumina公司的TruSeq RNA样本制备试剂盒构建链特异性cDNA文库：简言之，先将mRNA片段化，利用随机六聚体引物合成cDNA，并在第二链合成过程中掺入dUTP；接头连接完成后，使用尿嘧啶-N-糖基化酶去除第二链，随后进行PCR扩增。在Illumina NovaSeq 6000测序平台上开展双端测序。使用DESeq2进行差异表达分析，并通过美吉云（Majorbio Cloud）平台完成KEGG通路富集分析。