Adaptive RNA editing in Cephalopods

头足类动物的适应性 RNA 编辑

• 批准号: 22K15085
• 负责人: SANCHEZSILVA LUISGUSTAVO
• 金额: $ 2.91万
• 依托单位: Okinawa Institute of Science and Technology Graduate University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Early-Career Scientists
• 财政年份: 2022
• 资助国家: 日本
• 起止时间: 2022-04-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-22K15085/
• 关键词: RNA editing; Cephalopods; Single-cell sequencing; thermal adaptability

We have applied different single-cell RNA sequencing technologies in neural squid organs. One of these technologies uses a combination of 10X genomics and long-read sequencing with ONT or PacBio, from which we observed isoform-specific and RNA-editing at the 3' end corresponding to a cell type. However, the gene coverage and gene content per cell were still very low. We also observed that the highly repetitive genome of cephalopods limits the sequencing of entire gene isoforms and generates shorter sequences and significant levels of molecules coming from strand-invasion patterns. We have started sequencing single cells using plate-based sorting to solve our current limitations. Using our Euprymna berryi genome, I modified TSO-UMI primers to reduce the number of molecules with strand invasion. As a result, we have observed much larger cDNA peaks than in the previous technologies, indicating a successful fresh cell collection and skipping molecules with strand invasion during the RT-PCR.

我们在神经鱿鱼器官中应用了不同的单细胞RNA测序技术。这些技术之一使用10 X基因组学和ONT或PacBio的长读段测序的组合，从中我们观察到对应于细胞类型的3'端的亚型特异性和RNA编辑。然而，每个细胞的基因覆盖率和基因含量仍然很低。我们还观察到，头足类动物的高度重复的基因组限制了整个基因亚型的测序，并产生较短的序列和来自链入侵模式的分子的显着水平。 我们已经开始使用基于板的分选对单细胞进行测序，以解决我们目前的局限性。使用我们的Euprymna berryi基因组，我修改了TSO-UMI引物，以减少链侵入的分子数量。结果，我们观察到比先前技术中大得多的cDNA峰，表明成功的新鲜细胞收集和在RT-PCR期间具有链侵入的跳跃分子。