通过电穿孔对猪细胞系的CRISPR-Cas9基因组编辑的协议
Patricia Kiesler1, Stella S Lee2, Alexis L Norris2
1Biosystems and Biomaterials Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA.
STAR protocols
|October 11, 2024
概括
这项研究详细介绍了使用CRISPR-Cas9技术在猪丸细胞中进行基因组编辑的协议. 它涵盖了用于开发转基因动物的试剂评估,细胞输送和编辑评估.
科学领域:
- 动物生物技术动物生物技术
- 分子生物学分子生物学
- 基因组学就是基因组学.
背景情况:
- 基因组编辑为改善动物健康和公共卫生结果提供了潜力.
- 鉴定基因组编辑试剂和预测基因组变化的特征对于开发编辑动物至关重要.
- 猪丸 (ST) 细胞系是研究牲畜基因组编辑的相关模型.
研究的目的:
- 介绍猪丸 (ST) 细胞系中基因组编辑的详细方案.
- 建立一种标准化的方法来评估CRISPR-Cas9复杂功能.
- 促进基因编辑动物的开发,用于各种应用.
主要方法:
- 在体外评估CRISPR-Cas9复合物的活性.
- 通过转染,将基因组编辑分子输入ST细胞.
- 使用桑格测序对目标基因组变化的评估.
主要成果:
- 该协议为ST细胞中的基因组编辑提供了一种可复制的方法.
- 在体外成功评估了CRISPR-Cas9复杂的功能.
- 使用桑格测序证实了有针对性的编辑,证明了该协议的有效性.
结论:
- 这一协议使得在猪丸细胞中能够有效编辑基因组.
- 它是产生基因编辑动物的基础方法.
- 预计将在动物卫生和生物技术领域进一步应用.
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...


