Read Abigail, Gao Shaojian, Batchelor Eric, Luo Ji
Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD, USA.
Thoracic and Gastrointestinal Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, MD, USA.
Nucleic Acids Res. 2017 Jun 20;45(11):e101. doi: 10.1093/nar/gkx181.
CRISPR/Cas9-based gene knockout libraries have emerged as a powerful tool for functional screens. We present here a set of pre-designed human and mouse sgRNA sequences that are optimized for both high on-target potency and low off-target effect. To maximize the chance of target gene inactivation, sgRNAs were curated to target both 5΄ constitutive exons and exons that encode conserved protein domains. We describe here a robust and cost-effective method to construct multiple small sized CRISPR library from a single oligo pool generated by array synthesis using parallel oligonucleotide retrieval. Together, these resources provide a convenient means for individual labs to generate customized CRISPR libraries of variable size and coverage depth for functional genomics application.
基于CRISPR/Cas9的基因敲除文库已成为功能筛选的强大工具。我们在此展示了一组预先设计的人类和小鼠sgRNA序列，这些序列针对高靶向效力和低脱靶效应进行了优化。为了最大化靶基因失活的机会，sgRNA被精心设计以靶向5΄组成型外显子和编码保守蛋白结构域的外显子。我们在此描述了一种稳健且经济高效的方法，可从通过平行寡核苷酸检索的阵列合成产生的单个寡核苷酸池中构建多个小型CRISPR文库。总之，这些资源为各个实验室提供了一种便捷的方式，以生成大小和覆盖深度可变的定制CRISPR文库用于功能基因组学应用。
