一种随机多重CRISPRi-Seq方法用于识别基因的关键组合
Nicole A Ellis1, Kevin S Myers2,3, Jessica Tung1
1Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, United States.
eLife
|December 14, 2023
概括
一种新的多重,随机的CRISPR干扰测序 (MuRCiS) 方法快速识别病原体中的基本基因组合. 这种方法揭示了Legionella pneumophila中冗余的毒性基因,这对于宿主-病原体相互作用至关重要.
科学领域:
- 微生物学和遗传学 微生物学和遗传学
- 病原体的毒性机制病原体的毒性机制
- 基于CRISPR的基因查方法
背景情况:
- 病原体基因之间的功能冗余使得识别毒性关键因素变得复杂.
- 了解基因组合对于剖析复杂的生物结果至关重要.
- 现有的方法在快速查询大量冗余基因时是有限的.
研究的目的:
- 开发一种新的方法,多重,随机的CRISPR干扰测序 (MuRCiS),用于对基因组合的快速遗传查询.
- 在Legionella pneumophila*中识别病毒性关键基因组合.
- 探索MuRCiS对研究病变发生过程中的基因冗余的实用性.
主要方法:
- 开发了一种随机自组合方法,用于从合成寡核酸对中获得CRISPR阵列.
- 我们将其与PacBio的长读测序集成在一起,以进行全面的对对基因组合分析.
- 将MuRCiS应用于阿米巴和人类巨细胞模型中的44个Legionella pneumophila*毒性基因组.
主要成果:
- 成功调查了44个L.pneumophila*毒性基因的所有对联组合.
- 在阿米巴和人类巨模型中确定了几种对病原发生至关重要的基因的新组合.
- 发现的*lpg2888*和*lpg3000*在人类巨细胞中表现出冗余的作用,而*lpg3000*对于阿米巴病毒性至关重要.
结论:
- MuRCiS能够快速对大量冗余基因进行基因检查.
- 该方法有效地揭示了致病原体毒性所必需的基因组合.
- MuRCiS对各种生物系统和生物体的遗传研究具有广泛的适用性.
相关概念视频
CRISPR
Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced Short...
CRISPR and crRNAs
Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...
CRISPR
Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced Short...
CRISPR/Cas9 Genome Editing
The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...


