Developmental and Regenerative Biology Program, School of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong Special Administrative Region, China.
Department of Obstetrics and Gynaecology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, N.T., Hong Kong Special Administrative Region, China.
Int J Biochem Cell Biol. 2019 Feb;107:1-5. doi: 10.1016/j.biocel.2018.12.003. Epub 2018 Dec 6.
Single-cell parallel sequencing allows us to explore how genetic and epigenetic variations correlate of gene expression in the same cell. Beads-based approach and non-beads-based approach are the two present methods to separate DNA and RNA from the same cell. However, systematic difference between the two methods are lacking. In our study, we compared the performances of the two methods using transcriptome and methylome profiles generated simultaneously from single mouse oocytes. Our results showed that the beads-based approach could capture maximum quantity of mRNA but loss of DNA was inevitable, while the non-beads-based approach could obtain more DNA due to the undamaged nucleus obtained but at a cost of partial loss of mRNA. As the sequencing coverage of methylome sequencing in a single cell was relatively low, single-cell whole genome bisulfite sequencing (scWGBS) was preferable to generate the methylome map in single-cell parallel sequencing in comparison to single-cell reduced representation bisulfite sequencing (scRRBS). To the best of our knowledge, this is the first study to compare the two methods of single-cell parallel sequencing which offers a basic idea for deciding between the two methods and a direction of single-cell parallel sequencing development.
单细胞平行测序使我们能够探索基因和表观遗传变异如何与同一细胞中的基因表达相关。基于珠粒的方法和非基于珠粒的方法是目前从同一细胞中分离 DNA 和 RNA 的两种方法。然而，这两种方法之间缺乏系统差异。在我们的研究中，我们使用同时从单个小鼠卵母细胞中生成的转录组和甲基组谱来比较这两种方法的性能。我们的结果表明，基于珠粒的方法可以捕获最大量的 mRNA，但不可避免地会丢失 DNA，而基于非珠粒的方法由于获得了未受损的核，可以获得更多的 DNA，但代价是部分丢失了 mRNA。由于单个细胞中甲基组测序的测序覆盖率相对较低，与单细胞简化代表性亚硫酸氢盐测序（scRRBS）相比，单细胞全基因组亚硫酸氢盐测序（scWGBS）更适合在单细胞平行测序中生成甲基组图谱。据我们所知，这是首次比较单细胞平行测序的两种方法，为在这两种方法之间做出决定提供了基本思路，并为单细胞平行测序的发展指明了方向。
