基于染色体和等离子体的CRISPRi平台用于乳球菌乳糖中的条件基因沉默
Chenxi Huang1, Meishan Liu2, Jan Kok2
1Key Laboratory of Combinatorial Biosynthesis and Drug Discovery (Ministry of Education), School of Pharmaceutical Sciences, Wuhan University, Wuhan 430071, China.
International journal of molecular sciences
|October 16, 2025
概括
我们在乳球菌中开发了可诱导的CRISPR干扰 (CRISPRi) 系统,用于基因沉默. 基于染色体的系统为研究重要基因提供了严格的监管.
科学领域:
- 微生物学 微生物学
- 分子生物学分子生物学
- 基因组学就是基因组学.
背景情况:
- 克里斯普尔干扰 (CRISPRi) 是基因调节的一个强大工具.
- 在乳球菌中开发可诱导系统对于功能基因组学至关重要.
研究的目的:
- 建立和比较基于等离子体和基于染色体的可诱导CRISPRi系统在乳球菌中.
- 评估这些基因沉默系统的效率和调节.
主要方法:
- 使用缺核酶的Cas9 (dCas9) 和sgRNAs构建可诱导的CRISPRi系统.
- 使用了尼素诱导性 (PnisA) 和构成性 (Pusp45) 促进剂.
- 将dCas9集成到染色体中以改善调节.
- 执行了记者基因AcmA和基本基因ybeY的基因沉默.
- 对ybeY沉默细胞进行了转录组分析.
主要成果:
- 塑体和染色体系统都实现了基因抑制.
- 等离子体系统表现出泄漏的表达,而染色体集成确保了严格的调节.
- 基于染色体的CRISPRi (cbCRISPRi) 成功地使必要的基因ybeY沉默,导致生长缺陷.
- 转录组分析揭示了对核糖体蛋白,膜蛋白和转运体的显著影响.
结论:
- 可诱导的CRISPRi系统,特别是基于染色体的平台,为L. lactis的基因功能研究提供了强大的工具.
- 这些系统使得研究其他难以研究的基本基因的研究成为可能.
- cbCRISPRi系统为L. lactis的功能基因组学研究提供了精确的控制.
相关概念视频
01:25Inducible Operons: lac Operon
1.4K
The lac operon in Escherichia coli is a model for understanding inducible gene regulation and metabolic flexibility. It integrates local control by lactose and global regulation through catabolite repression, enabling E. coli to preferentially metabolize glucose when available and switch to lactose utilization when glucose is scarce.Structure and Function of the lac OperonThe lac operon contains three structural genes: lacZ (β-galactosidase), lacY (lactose permease), and lacA...
1.4K
02:53Conservative Site-specific Recombination and Phase Variation
6.6K
Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...
The recognition sites for Cre recombinase called LoxP...
6.6K
01:59CRISPR
57.5K
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...
57.5K
01:58Prokaryotic Transcriptional Activators and Repressors
25.1K
The organization of prokaryotic genes in their genome is notably different from that of eukaryotes. Prokaryotic genes are organized, such that the genes for proteins involved in the same biochemical process or function are located together in groups. This group of genes, along with their regulatory elements, are collectively known as an operon. The functional genes in an operon are transcribed together to give a single strand of mRNA known as polycistronic mRNA.
Transcription of prokaryotic...
Transcription of prokaryotic...
25.1K
02:53CRISPR and crRNAs
18.7K
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...
18.7K