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FAQ
How is Prime Editing 7 (PE7) different from traditional CRISPR/Cas9 technology?
Traditional CRISPR/Cas9 technology achieves gene editing by introducing double-strand breaks at the target DNA site and then using the cell’s homologous recombination repair mechanism. This approach carries multiple risks, such as lower editing efficiency, reduced homozygous mutation rates, and random insertions or deletions. Prime Editing, however, does not require double-strand breaks. With its Cas9n-RT editing enzyme system and pegRNA, Prime Editing achieves more accurate and safer gene editing with reduced off-target effects.
Are all types of genes suitable for KO cell lines?
Not all genes are suitable for knockout. Some gene knockouts may result in cell death or severe dysfunction, particularly for essential genes. In such cases, conditional knockouts or gene knockdowns (e.g., RNAi) may be used instead.
Why choose EDITGENE?
EDITGENE provides access to a comprehensive library of over 4,500 high-quality knockout (KO) cell lines, enabling researchers to save valuable time. Our custom gene knockout services are highly efficient, boasting a high positive rate while minimizing off-target effects. Clients also benefit from personalized, one-on-one support from a team of PhD experts from globally renowned institutions, ensuring top-tier service and results.
What is the difference between KO cell lines and gene knockout animal models?
KO cell lines are used for in vitro experiments, suitable for high-throughput screening and cellular studies, while gene knockout animal models are used for in vivo experiments to study gene functions within an entire organism and its interaction with the environment.
Why do researchers use KO cell lines?
Researchers use KO cell lines to investigate gene functions by observing the effects of gene deletion on cellular behavior. This helps in understanding the role of genes in various processes like cell growth, metabolism, and signal transduction. KO cell lines are vital for studying diseases like cancer, genetic disorders, and neurodegenerative diseases.
What is a KO cell line?
KO (Knockout) cell line is a cell line where a specific gene has been completely removed or rendered non-functional through gene editing technologies such as CRISPR-Cas9. These cell lines are critical for understanding gene functions and disease mechanisms.
Are KO cell lines applicable to all cell types?
KO cell lines can be applied to various cell types, including cancer cells, stem cells, and primary cells, but different cell types may have varying sensitivities to gene editing, and may vary among different cell types. In certain cell types, achieving gene knockout may require optimization of transfection conditions and selection of appropriate gene-editing tools.
What is gene overexpression?
Gene overexpression refers to using various techniques to significantly increase the expression level of a specific gene in cells or organisms. This is often achieved by introducing additional gene copies or using strong promoters to drive gene expression.
Why conduct gene overexpression?
Gene overexpression aids in studying the function of specific genes, revealing their role within the organism. It is also commonly used in drug screening, vaccine development, and protein production. For example, by overexpressing a therapeutic protein, researchers can evaluate its efficacy in disease models.
KO细胞系是否适合所有类型的基因？
虽然KO细胞系技术强大，但并非所有基因都适合敲除。某些基因的敲除可能导致细胞死亡或严重的生理功能障碍，特别是那些对细胞存活至关重要的基因。在这种情况下，可以选择条件性敲除或基因敲降（如RNAi）来研究这些基因的功能。
什么是KO细胞系？
KO细胞系（Knockout Cell Line）是顺利获得基因编辑技术（如CRISPR-Cas9）敲除或删除特定基因的细胞系。这意味着目标基因被完全移除或失去功能，使研究人员能够观察该基因缺失对细胞行为和生物学过程的影响。这种细胞系在生物医学研究中非常重要，因为它们给予了直接分析基因功能和疾病机制的实验工具。
一般CRISPR相关试剂以及Cas蛋白可以存放多久？
CRISPR检测相关试剂：
①RPA恒温扩增试剂盒于-20℃保存，可长期存放。
②靶标质粒可长期存放-20℃ ③ Cas蛋白反复冻融容易影响活性，建议分装多管，存放-80℃，根据实验需要取出使用。另外，短期内使用的，可放-20℃保存。
③crRNA容易降解，建议短时间内用不到的于-80℃保存。
④探针是DNA双链，相对来说没有那么容易降解，可放-20℃保存。
①RPA恒温扩增试剂盒于-20℃保存，可长期存放。
②靶标质粒可长期存放-20℃ ③ Cas蛋白反复冻融容易影响活性，建议分装多管，存放-80℃，根据实验需要取出使用。另外，短期内使用的，可放-20℃保存。
③crRNA容易降解，建议短时间内用不到的于-80℃保存。
④探针是DNA双链，相对来说没有那么容易降解，可放-20℃保存。
为什么要进行基因过表达？
基因过表达可以帮助研究特定基因的功能，揭示其在生物体内的作用机制。它也常用于药物筛选、疫苗开发和蛋白质生产等场景中。例如，顺利获得过表达某个治疗蛋白，可以研究其在疾病模型中的治疗效果。
基因敲入在药物开发中的作用是什么?
基因敲入在药物开发中发挥了重要作用。它用于靶点验证，顺利获得将特定基因敲入细胞系或动物模型中，确认药物靶点的有效性。它还能帮助建立疾病模型，测试药物在这些模型上的疗效和安全性。基因敲入技术还支持药物筛选，顺利获得带有敲入基因的细胞系进行高通量筛选，发现潜在药物候选分子。此外，它还能揭示药物的作用机制，优化药物结构和用法。整体而言，基因敲入技术加速了药物研发过程，并提高了治疗效果和安全性。
如何选择合适的细胞进行文库筛选？
细胞选择可以遵循以下原则：
1）与研究目的相符
2）sgRNA文库靶向的基因要与细胞的属源相符
3）细胞可以稳定传代
4）转染效率高
尽量不选用原代细胞。由于原代细胞无法稳定传代，可能在文库筛选实验的过程中，原代细胞已经出现大量死亡，无法完成实验。如果选用原代细胞进行文库筛选，为分析决上述的风险，可以顺利获得降低细胞覆盖度和选择gRNA数较小的文库进行筛选，尽可能降低文库细胞池的数量和缩短实验周期。
1）与研究目的相符
2）sgRNA文库靶向的基因要与细胞的属源相符
3）细胞可以稳定传代
4）转染效率高
尽量不选用原代细胞。由于原代细胞无法稳定传代，可能在文库筛选实验的过程中，原代细胞已经出现大量死亡，无法完成实验。如果选用原代细胞进行文库筛选，为分析决上述的风险，可以顺利获得降低细胞覆盖度和选择gRNA数较小的文库进行筛选，尽可能降低文库细胞池的数量和缩短实验周期。
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