Copy number variation of genes (exons) in invasive Cottus compared to the parental species, Cottus rhenanus and Cottus perifretum.

Hybridization between Cottus rhenanus and C. perifretum has resulted in an evolutionary young hybrid lineage of invasive Cottus that has colonized a new habitat where the parental species are not found (Nolte et al. 2005; Proc. R. Soc. B 272: 2379–2387). This CGH array was designed to screen for copy number variation among Cottus species and to find gene duplicates that are unique to the hybrid lineage (see also Dennenmoser et al. 2017; Copy number increases of transposable elements and protein coding genes in an invasive fish of hybrid origin). SurePrint G3 custom CGH 8x60k microarray with five technical replicates per probe and a 15 bp linker added to each probe as recommended by Agilent. The experimental design was a two-color experiment (Cottus individual vs. common reference) with 24 biological replicates (3 species x 8 individuals). Each Cottus individual was co-hybridized with the same Cottus perifretum individual as a common reference. The normalization procedure included a calibration step to account for probe-specific binding behavior, which was assessed using a custom designed calibration array (see also Dennenmoser et al. 2017; Copy number increases of transposable elements and protein coding genes in an invasive fish of hybrid origin).

莱茵杜父鱼（Cottus rhenanus）与佩氏杜父鱼（Cottus perifretum）的杂交事件，催生了一个进化历程较短的入侵性杜父鱼杂交谱系，该谱系已占据亲本物种未曾涉足的新生境（Nolte等，2005；《英国皇家学会学报B辑》，272卷：2379–2387）。本研究设计的比较基因组杂交（Comparative Genomic Hybridization, CGH）芯片，旨在筛选杜父鱼物种间的拷贝数变异，并鉴定该杂交谱系特有的基因重复事件（另见Dennenmoser等，2017；《杂交起源入侵鱼类中转座元件与蛋白编码基因的拷贝数扩增》）。该芯片为安捷伦（Agilent）定制的SurePrint G3型8×60K微阵列，每个探针设置5次技术重复，并按照安捷伦的推荐方案为每个探针添加15 bp的接头序列。本实验采用双色杂交设计（杜父鱼个体 vs 通用参考样本），共设置24个生物学重复样本（3个物种×8个个体），每个杜父鱼个体均与同一尾佩氏杜父鱼个体作为通用参考样本进行共杂交。归一化流程包含校准步骤，以校正探针特异性结合行为，该校准通过定制校准芯片完成（另见Dennenmoser等，2017；《杂交起源入侵鱼类中转座元件与蛋白编码基因的拷贝数扩增》）。