全长的异形连接测序来解决癌症转录组复杂性的问题
Saranga Wijeratne1, Maria E Hernandez Gonzalez1, Kelli Roach1
1The Steve and Cindy Rasmussen Institute for Genomic Medicine, Abigail Wexner Research Institute at Nationwide Children's Hospital, 575 Children's Crossroad, Columbus, OH, 43215, USA.
BMC genomics
|January 29, 2024
概括
我们开发了PacBio全长异形并列体测序 (PB_FLIC-Seq),以增加长读测序输出,改善新型癌症异形的检测. 这种方法增强了儿科质瘤中的异型发现,揭示了显著的细胞外基因表达变化.
科学领域:
- 基因组学就是基因组学.
- 分子生物学分子生物学
- 癌症研究 癌症研究
背景情况:
- 癌症转录组是复杂的,异常拼接导致差异化异型表达.
- 短读测序在准确推断全长转录时存在局限性.
- 长读数测序 (Iso-Seq) 提供全长的异形分辨率,但读数输出较低.
研究的目的:
- 开发一种新的连接工作流程,即PacBio全长异形连接序列测序 (PB_FLIC-Seq),以增加独特的长读数.
- 为了提高中度到低水平表达异型体的检测,并改善整体异型体发现.
- 将PB_FLIC-Seq应用于癌症转录组的分析,特别是儿科扩散中线质瘤.
主要方法:
- 开发并实施了PB_FLIC-Seq连接工作流程,用于PacBio长读RNA测序.
- 测序了一个商业参考 (Spike-In RNA变种; SIRV) 来评估方法的性能.
- 应用PB_FLIC-Seq来分析儿科扩散性中线质瘤和相邻的非恶性组织.
主要成果:
- 与标准的Iso-Seq相比,PB_FLIC-Seq显示出读输出增加了3.4倍,并改善了SIRV回忆.
- 在质瘤中确定了差异性全长异构体表达,包括分泌蛋白酸和氨酸丰富异构体 (SPARC) 的11676倍增加.
- 使用PB_FLIC-Seq方法检测到几种新的癌症特异性异构体.
结论:
- 该PB_FLIC-Seq方法有效地增加了PacBio平台上测序的全长异型的读数.
- 这种方法可以更好地发现表达的异型,这对于复杂的转录组表征至关重要.
- 在儿科质瘤中PB_FLIC-Seq的应用突出了其在识别癌症特异性异型表达中的实用性.
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