Analysis and comprehensive comparison of Pacbio and Nanopore-based RNA-sequencing in Arabidopsis transcriptome. Analysis and comprehensive comparison of Pacbio and Nanopore-based RNA-sequencing in Arabidopsis transcriptome

Rapidly increased studies by third-generation sequencing [Pacific Biosciences (Pacbio) and Oxford Nanopore Technologies (ONT)] have been used in all kinds of research areas. Among them, the plant full-length single-molecule transcriptome studies were most used by Pacbio while ONT was rarely used. Therefore, in this study, we developed ONT RNA-sequencing methods in plants. We performed a detailed evaluation of reads from Pacbio and Nanopore PCR cDNA (ONT Pc) sequencing in plants (Arabidopsis), including the characteristics of raw data and identification of transcripts. We aimed to provide a valuable reference for applications of ONT in plant transcriptome analysis. Overall design: Sequencing the Arabidopsis cDNA samples from overall ground parts at the same growth stage by Pacbio and Nanopore platform repectively. And then, we compared the advantages and disadvantages between Pacbio and Nanopore RNA-seq. Please note that the Sample 1 RNA-seq is to generate a transcriptome assembly and the rest of the samples are to quantify the gene expression.

第三代测序技术（third-generation sequencing）涵盖Pacific Biosciences（Pacbio）与Oxford Nanopore Technologies（ONT）两大平台，相关研究数量快速增长，现已应用于各类研究领域。其中，Pacbio平台最常用于植物全长单分子转录组研究，而ONT的应用则相对有限。为此，本研究针对植物样本开发了ONT RNA测序方法。我们以拟南芥（Arabidopsis）为实验材料，对Pacbio与Nanopore PCR cDNA（ONT Pc）测序所得的读段（reads）进行了详细评估，内容涵盖原始数据特征分析与转录本鉴定两大环节，旨在为ONT在植物转录组分析中的应用提供有价值的参考依据。
整体实验设计：分别采用Pacbio与Nanopore测序平台，对同一生长阶段拟南芥全地上部分的cDNA样本进行测序；随后对比两种平台在RNA测序（RNA-seq）应用中的优劣势。请注意，样本1的RNA-seq用于构建转录本组装集，其余样本则用于基因表达定量分析。