Build your own CRISPR solution

The powerful CRISPR gene editing technology transforms research at an astonishing rate. Read about a five-step approach for DIY CRISPR workflow, with focus on CRISPR-Cas9 vector design, its construction and delivery into cells, possible ways for mutant genotyping and characterization. We’ve refined and optimized each step to help ensure maximum editing efficiency across a broad spectrum of cell types.

The CRISPR-Cas9 system is a two-component system, consisting of the target-specific CRISPR gRNA and Cas9 nuclease. For genome editing to be successful, both the Cas9 and gRNA need to be expressed together in the target cells. We offer the necessary tools for the design and engineering of target-specific gRNA and Cas9 nuclease expression plasmids enabling you to pursue different experimental strategies.

Products for CRISPR-Cas9 design

For designing your vector you may chose between two separate plasmids for Cas9 and gRNA, respectively, or a single plasmid for both genes:

  • Plasmid for Cas nuclease expression obtained from open-access resources (e.g. Addgene, the nonprofit global plasmid repository)
  • gRNA cloning vector obtained from open-access resources (e.g. Addgene, the nonprofit global plasmid repository)
  • GeneArt CRISPR Nuclease vector for cloning Cas9 and gRNA together

For constructing your own plasmid for gRNA expression you may need:

For easy cloning of novel CRISPR nuclease sequences or subcloning work you may need:

Anza Restriction Enzymes

Invitrogen TrueTag Donor DNA Kits can help you obtain up-to 100% knocked in cells using our predesigned and validated donor DNA templates.

Looking for our premium genome editing solutions? Learn more about our first-class genome engineering products, including our TrueCut Cas9 v2 and TrueGuide Synthetic sgRNA.

Learn more

Once DNA fragments coding for CRISPR-Cas9 system components are cloned, the vector can be propagated within E. coli cells to generate sufficient quantities of your expression plasmid. We offer a variety of competent E. coli cells, selection of which depends upon the transformation method, and throughput of your experiment.

There are different methods to verify that your plasmid construct is correct, i.e. PCR, restriction digestion or sequencing. The choice depends on whether you are interested in determining if the plasmid contains your DNA insert, is the insert in the right orientation, or does the insert have the correct sequence. We provide molecular biology tools to implement any analysis method you may choose.

Products used for plasmid transformation and subsequent screening of clones

For efficient transformation of constructed CRISPR plasmids you need competent E. coli cells. We recommend using One Shot MAX Efficiency DH5a T1R Chemically Competent Cells (additional products and sizes are available).

Isolate plasmid DNA in sufficient quantities and maximal purity for subsequent delivery to eukaryotic cells. Use PureLink Expi Endotoxin-Free Maxi Plasmid Purification Kit to generate high yields of endotoxin-free plasmid (additional products and sizes are available).

Before proceeding into following steps you may wish to verify that your plasmid constructs are correct. Chose between different analysis methods:

Accessory products:

dh5a competent cells

The CRISPR-Cas9 system greatly simplifies genome editing and has great promise in broad applications such as stem cell engineering, gene therapy, tissue and animal disease models, and engineering disease-resistant transgenic plants.

Transfection is the process by which CRISPR-Cas9 DNA, mRNA, or protein systems are introduced into eukaryotic cells. Construct delivery techniques vary widely and include lipid nanoparticle–mediated transfection, viral delivery, and physical methods such as electroporation.

Products used for CRISPR construct delivery into eukaryotic cells

Cell Culture Tools and Essentials:

  • Sera - for your specific cell culture needs—from basic research to specialty assays
  • Cell Culture Plastics - for optimum cell growth and consistency in cell culture
  • Cell Culture Essentials - find the tools and resources you need for successful cell culture
Lipofectamine

Whichever genome editing strategy you use, careful monitoring of the process will help you generate robust and reliable results. Start with accurate cell counts and viability determinations, then screen and validate the genotype of your cells. The genotyping technique of the mutant sequence depends on the type of mutation introduced through the CRISPR-generated edit. Here are the most common techniques:

  • PCR amplification and gel electrophoresis for detection of larger Indels
  • Mismatch-cleavage assay for Indel detection (T7 Endonuclease I cleavage assay)
  • PCR amplification and restriction digestion for HDR analysis
  • PCR amplification and cloning followed by Sanger sequencing
  • PCR amplification and NGS

Products used to detect CRISPR-mediated genome modifications

Zero Blunt TOPO PCR Cloning Kit for Sequencing

CRISPR is routinely used for knockout, knock-in, or modulation of gene expression, and the effects can be measured using cell analysis techniques. Real-time PCR allows monitoring changes in the expression at gene level, for example when non-sense mediated decay decreases transcript levels, while western blotting is used to view changes to protein expression in a cell population; flow cytometry provides the throughput for multiparameter analysis on vast numbers of individual cells. Imaging allows for direct analysis of changes in protein expression, compartmentalization, and cell morphology, while high-content analysis (HCA) provides automation for the imaging process with quantitative rigor.

Products used for further CRISPR analysis and edited cell collection

UltraPure DNase RNase

The CRISPR-Cas9 system is a two-component system, consisting of the target-specific CRISPR gRNA and Cas9 nuclease. For genome editing to be successful, both the Cas9 and gRNA need to be expressed together in the target cells. We offer the necessary tools for the design and engineering of target-specific gRNA and Cas9 nuclease expression plasmids enabling you to pursue different experimental strategies.

Products for CRISPR-Cas9 design

For designing your vector you may chose between two separate plasmids for Cas9 and gRNA, respectively, or a single plasmid for both genes:

  • Plasmid for Cas nuclease expression obtained from open-access resources (e.g. Addgene, the nonprofit global plasmid repository)
  • gRNA cloning vector obtained from open-access resources (e.g. Addgene, the nonprofit global plasmid repository)
  • GeneArt CRISPR Nuclease vector for cloning Cas9 and gRNA together

For constructing your own plasmid for gRNA expression you may need:

For easy cloning of novel CRISPR nuclease sequences or subcloning work you may need:

Anza Restriction Enzymes

Invitrogen TrueTag Donor DNA Kits can help you obtain up-to 100% knocked in cells using our predesigned and validated donor DNA templates.

Looking for our premium genome editing solutions? Learn more about our first-class genome engineering products, including our TrueCut Cas9 v2 and TrueGuide Synthetic sgRNA.

Learn more

Once DNA fragments coding for CRISPR-Cas9 system components are cloned, the vector can be propagated within E. coli cells to generate sufficient quantities of your expression plasmid. We offer a variety of competent E. coli cells, selection of which depends upon the transformation method, and throughput of your experiment.

There are different methods to verify that your plasmid construct is correct, i.e. PCR, restriction digestion or sequencing. The choice depends on whether you are interested in determining if the plasmid contains your DNA insert, is the insert in the right orientation, or does the insert have the correct sequence. We provide molecular biology tools to implement any analysis method you may choose.

Products used for plasmid transformation and subsequent screening of clones

For efficient transformation of constructed CRISPR plasmids you need competent E. coli cells. We recommend using One Shot MAX Efficiency DH5a T1R Chemically Competent Cells (additional products and sizes are available).

Isolate plasmid DNA in sufficient quantities and maximal purity for subsequent delivery to eukaryotic cells. Use PureLink Expi Endotoxin-Free Maxi Plasmid Purification Kit to generate high yields of endotoxin-free plasmid (additional products and sizes are available).

Before proceeding into following steps you may wish to verify that your plasmid constructs are correct. Chose between different analysis methods:

Accessory products:

dh5a competent cells

The CRISPR-Cas9 system greatly simplifies genome editing and has great promise in broad applications such as stem cell engineering, gene therapy, tissue and animal disease models, and engineering disease-resistant transgenic plants.

Transfection is the process by which CRISPR-Cas9 DNA, mRNA, or protein systems are introduced into eukaryotic cells. Construct delivery techniques vary widely and include lipid nanoparticle–mediated transfection, viral delivery, and physical methods such as electroporation.

Products used for CRISPR construct delivery into eukaryotic cells

Cell Culture Tools and Essentials:

  • Sera - for your specific cell culture needs—from basic research to specialty assays
  • Cell Culture Plastics - for optimum cell growth and consistency in cell culture
  • Cell Culture Essentials - find the tools and resources you need for successful cell culture
Lipofectamine

Whichever genome editing strategy you use, careful monitoring of the process will help you generate robust and reliable results. Start with accurate cell counts and viability determinations, then screen and validate the genotype of your cells. The genotyping technique of the mutant sequence depends on the type of mutation introduced through the CRISPR-generated edit. Here are the most common techniques:

  • PCR amplification and gel electrophoresis for detection of larger Indels
  • Mismatch-cleavage assay for Indel detection (T7 Endonuclease I cleavage assay)
  • PCR amplification and restriction digestion for HDR analysis
  • PCR amplification and cloning followed by Sanger sequencing
  • PCR amplification and NGS

Products used to detect CRISPR-mediated genome modifications

Zero Blunt TOPO PCR Cloning Kit for Sequencing

CRISPR is routinely used for knockout, knock-in, or modulation of gene expression, and the effects can be measured using cell analysis techniques. Real-time PCR allows monitoring changes in the expression at gene level, for example when non-sense mediated decay decreases transcript levels, while western blotting is used to view changes to protein expression in a cell population; flow cytometry provides the throughput for multiparameter analysis on vast numbers of individual cells. Imaging allows for direct analysis of changes in protein expression, compartmentalization, and cell morphology, while high-content analysis (HCA) provides automation for the imaging process with quantitative rigor.

Products used for further CRISPR analysis and edited cell collection

UltraPure DNase RNase