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从环境微核细胞中获得ASV解析的rRNA操作子的长读测序策略的基准测试
Christina Karmisholt Overgaard1, Mahwash Jamy2,3, Simona Radutoiu4
1Center for Microbial Communities, Department of Chemistry and Bioscience, Aalborg University, Aalborg, Denmark.
Molecular ecology resources
|July 9, 2024
概括
创建一个合成真核生物模拟社区使得长期阅读的测序和生物信息学工具用于微核生物分类的评估成为可能. 这推动了18S rRNA基因数据库的开发和环境DNA的分析.
科学领域:
- 环境微生物学环境微生物学
- 分子生态学分子生态学
- 生物信息学是一种生物信息学.
背景情况:
- 微核细胞分类的短读元编码受到不完整的18S rRNA基因参考数据库的限制.
- 长读测序提供了扩大类遗传覆盖范围的潜力,但需要进行性能评估.
- 缺乏明确定义的真核生物模拟社区,阻碍了对测序和生物信息学工具的评估.
研究的目的:
- 开发一个精确定义的真核体模拟社区,用于评估测序和生物信息学性能.
- 评估三种长读序列策略和三种幅序列变异 (ASV) 调用工具.
- 为具有成本效益,高通量rRNA操作子测序提供建议.
主要方法:
- 构建一个真核rRNA操作子克隆库,以创建一个合成模拟社区.
- 评估PacBio循环共识测序和两个纳米孔测序方法.
- 评估USEARCH,VSEARCH和DADA2用于ASV分辨率,专注于奇默的检测.
主要成果:
- 确定了不同长读序列策略和ASV呼叫者的性能指标 (灵敏度和准确性).
- 该研究确定了获得高质量的rRNA操作子的最佳方法,以最小的错误.
- 模拟社区的结果在多样化的农业土壤样本中得到了验证,揭示了新的微核细胞系.
结论:
- 开发的真核细胞模拟社区是验证测序和生物信息学管道的宝贵资源.
- 建立了最佳实践协议,用于高通量,无错误的rRNA操作子测序.
- 这项工作有助于改善环境样本中的微核细胞识别和发现.
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