INRAE, Fish Physiology and Genomics, UR 1037, Rennes, France.
Department of Production Animal Clinical Sciences, Faculty of Veterinary Medicine, Norwegian University of Life Sciences, Ås, Norway.
Fish Physiol Biochem. 2024 Oct;50(5):2067-2082. doi: 10.1007/s10695-024-01326-5. Epub 2024 Mar 1.
DNA methylation in CpG dinucleotides is an important epigenetic mark in fish spermatozoa since it has been shown that some sperm methylome features are transmitted to the offspring. Reduced representation bisulfite sequencing (RRBS) is one genome-scale methods developed to assess DNA methylation at CpG sites. It allows the sequencing of a reduced fraction of the genome expected to be enriched in CpGs. The aim of this study is to characterize the extent of the CpG sites that can be identified in the RRBS-reduced sequenced fraction of rainbow trout spermatozoa, in order to evaluate the potential of RRBS for sperm DNA methylation studies. We observed that RRBS did provide a reduced amount of genomic data, the sum of the CpGs analyzed on 12 males spanning 9% of the total genomic CpGs. CpGs were only slightly enriched in the RRBS data (×1.7 times the sequenced nucleotides), the possible causes being linked to trout genome structure and sequenced fragments size. All genomic functional features were represented in our CpG dataset, with a noticeable enrichment in exons but, strikingly, not in promoters. The number of CpGs shared between biological replicates was low, but this proportion reached workable values from six biological replicates (46% of the analyzed cytosines) on. The choices that are to be made regarding fragment size selection and the options during bioinformatic data processing are discussed. In all, RRBS is a relevant first-approach method to scan the CpG DNA methylation status of spermatozoa along rainbow trout genome, although in a very reduced pattern among biological replicates.
DNA 甲基化在 CpG 二核苷酸中是鱼类精子中的一种重要的表观遗传标记，因为已经表明一些精子甲基组学特征会传递给后代。缩减代表性亚硫酸氢盐测序 (RRBS) 是一种用于评估 CpG 位点 DNA 甲基化的全基因组方法。它允许对预期富含 CpG 的基因组的缩减部分进行测序。本研究的目的是描述可以在虹鳟精子 RRBS 缩减测序部分中鉴定的 CpG 位点的程度，以评估 RRBS 在精子 DNA 甲基化研究中的潜力。我们观察到 RRBS 确实提供了较少的基因组数据，对 12 个雄性个体的 12%的基因组 CpG 进行了分析。CpG 在 RRBS 数据中仅略有富集（测序核苷酸的 1.7 倍），可能的原因与鳜鱼基因组结构和测序片段大小有关。我们的 CpG 数据集包含所有基因组功能特征，在外显子中明显富集，但令人惊讶的是，在启动子中没有富集。在生物学重复之间共享的 CpG 数量很少，但从六个生物学重复（分析的胞嘧啶的 46%）开始，这一比例达到了可行的值。讨论了在片段大小选择和生物信息学数据处理过程中的选项方面需要做出的选择。总之，RRBS 是一种相关的初步方法，可以扫描虹鳟鱼精子基因组中的 CpG DNA 甲基化状态，尽管在生物学重复中呈现出非常低的模式。
