CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) and CRISPR-associated (Cas) genes are essential in adaptive immunity in select bacteria and archaea, enabling the organisms to respond to and eliminate invading genetic material.
- Determining Genome Targeting Efficiency using T7 Endonuclease I
- EnGen® sgRNA Synthesis Kit, S. pyogenes Protocol (E3322)
- In vitro digestion of DNA with Cas9 Nuclease, S. pyogenes (M0386)
- NEBuilder HiFi DNA Assembly Reaction Protocol
- NEBuilder® HiFi DNA Assembly Chemical Transformation Protocol
- NEBuilder® HiFi DNA Assembly Chemical Transformation Protocol (E2621)
- NEBuilder® HiFi DNA Assembly Electrocompetent Transformation Protocol
- NEBuilder® HiFi Electrocompetent Transformation Protocol (E2621)
- PCR Using Q5® Hot Start High-Fidelity DNA Polymerase (M0493)
- sgRNA Synthesis Using the HiScribe™ Quick T7 High Yield RNA Synthesis Kit (NEB #E2050)
- T7 Endonuclease I-based Mutation Detection with the EnGen® Mutation Detection Kit (NEB #E3321)
- Transfection of Cas9 RNP (ribonucleoprotein) into adherent cells using the Lipofectamine® RNAiMAX
- Using recombinant Cas9 nuclease to assess locus modification in genome editing experiments (#M0386)
CRISPR/Cas9 & Targeted Genome Editing: New Era in Molecular Biology
Understand the history, importance and future of CRISPR/Cas9 and target genome editing
- Genome Editing Brochure
- Kinetic Comparison of Cas9 Homologs Recognizing Diverse PAM Sequences (2018)
- Zheng Hu, Wencheng Ding, Da Zhu, Lan Yu, Xiaohui Jiang, Xiaoli Wang, Changlin Zhang, Liming Wang, Teng Ji, Dan Liu, Dan He, Xi Xia, Tao Zhu, Juncheng Wei, Peng Wu, Changyu Wang, Ling Xi, Qinglei Gao, Gang Chen, Rong Liu, Kezhen Li, Shuang Li, Shixuan Wang, Jianfeng Zhou, Ding Ma, Hui Wang 2015. TALEN-mediated targeting of HPV oncogenes ameliorates HPV-related cervical malignancy. J Clin Invest. 125, PubMedID: 25500889, DOI: 10.1172/JCI78206
- A-Bk Dad, S Ramakrishna, M Song, H Kim 2014. Enhanced gene disruption by programmable nucleases delivered by a minicircle vector. Gene Ther. , PubMedID: 25142139, DOI: 10.1038/gt.2014.76
Online ResourcesPlasmid Repositories:
CRISPR-gRNA Design Tools:
Genome Engineering using CRISPR/Cas Systems
Cas9-triggered homologous recombination
Drosophila RNAi Screening Center at Harvard Medical School
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Get a high level overview of CRISPR/Cas9 and how it is used in genome editing in our Science in 60 segment.
CRISPR systems evolved as a means of bacterial adaptive immunity. Understanding their behavior in vivo is essential to harnessing their power in vitro.
In this webinar you will learn how to increase editing efficiency by directly introducing Cas9 ribonucleoproteins (RNPs) to cells through electroporation or lipofection. Rapid sgRNA synthesis requiring only a single user-supplied ~55mer single-stranded DNA oligonucleotide is described. Methods for assessing genome editing efficiency will be discussed including T7-endonuclease I-based methods, sequencing-based methods, and in vitro Cas9 digestion.