Transcriptome of ΔPfPRMT5 parasite during malaria parasite asexual development [PRMT5_RNAseq]. Transcriptome of ΔPfPRMT5 parasite during malaria parasite asexual development [PRMT5_RNAseq]

Purpose: this study is to analyze the change of RNA splicing events after disruption of Protein Arginine Methyltranferase 5 (PRMT5) in Plasmodium falciparum. Methods: In this study, the transcriptomes of a PfPRMT5 gene knockout (KO) parasite line with its wildtype control were analyzed by RNAseq.Total RNA were harvested from the asexual parasites at four stages (ring, early trophozoite, late trophozoite, and schizont)(12h, 24h, 36h, and 46h post-invasion) using the Quick-RNA MiniPrep kit (Zymo Research). RNA sequencing libraries were prepared using the KAPA stranded RNA-seq library preparation kit (Roche) with 500 ng RNA from each sample. Illumina adapter sequence removal and quality trimming of reads were performed using Trimmomatic. Only reads that had a minimum length of 50 base pairs were retained. Reads were then mapped to the P. falciparum 3D7 strain reference genome with HISAT2. Results: This RNAseq analysis with strand-specific mRNA libraries from both ΔPfPRMT5 and WT lines showed that >90% of sequencing reads were of high quality for mapping to the P. falciparum genome with nearly 40 times of coverage. This allows us to analyze the change of alternative splicing events (alternative 5' splice site, alternative 3' splice site, retained intron and skipped exon). 800, 1056, 479, and 1158 alternative splicing events (alternative 5' splice site, alternative 3' splice site, retained intron and skipped exon) were altered in the DPfPRMT5 parasite line as compared to the WT line at four development stages, respectively (unpublished data). Conclusions: Collectively, this RNAseq provide a dataset for analysis of abnormal RNA splicing events in the ΔPfPRMT5 parasites. Overall design: Transcriptome of ΔpfPRMT5 parasites at four developmental stages for in-depth RNA splicing analysis

研究目的：本研究旨在分析恶性疟原虫（Plasmodium falciparum）中蛋白质精氨酸甲基转移酶5（Protein Arginine Methyltranferase 5, PRMT5）敲除后RNA剪接事件的变化。
研究方法：本研究通过RNA测序（RNAseq）分析了PfPRMT5基因敲除（KO）寄生虫株与其野生型对照的转录组。采用Quick-RNA MiniPrep试剂盒（Zymo Research公司），从入侵后12h、24h、36h、46h的环状体、早期滋养体、晚期滋养体、裂殖体四个无性发育阶段的寄生虫中提取总RNA。使用KAPA链特异性RNA-seq文库制备试剂盒（罗氏（Roche）公司），以每份样本500ng RNA构建测序文库。采用Trimmomatic工具去除Illumina接头序列并对测序reads进行质量修剪，仅保留长度不低于50碱基对的reads。随后通过HISAT2将reads比对至恶性疟原虫3D7株参考基因组。
研究结果：本研究针对ΔPfPRMT5与野生型（WT）株的链特异性mRNA文库开展RNA测序分析，结果显示超过90%的测序reads质量优良，可比对至恶性疟原虫基因组，覆盖度近40倍。这为我们分析可变剪接事件（可变5'剪接位点、可变3'剪接位点、内含子保留和外显子跳跃）的变化提供了条件。相较于野生型株，ΔPfPRMT5寄生虫株在四个发育阶段分别发生了800、1056、479和1158个可变剪接事件（对应可变5'剪接位点、可变3'剪接位点、内含子保留和外显子跳过）的改变（未发表数据）。
研究结论：综上，本RNA测序数据集可用于分析ΔPfPRMT5寄生虫中的异常RNA剪接事件。
整体实验设计：对ΔpfPRMT5寄生虫四个发育阶段的转录组进行测序，以开展深入的RNA剪接分析。