CRISPR Genome Editing Tools and Services

• Lentivector: spCas9 + sgRNA + Puro^R — the standard choice for stable knockout workflows.
• AAV: saCas9 + sgRNA — suited to size-constrained AAV delivery.
• Non-viral: spCas9 + sgRNA + GFP — suited to transfection-compatible cultured cells.

Design note: For size-limited viral constructs, keep the selectable marker or reporter configuration simple. AAV workflows commonly use saCas9 because it is smaller than spCas9.

• ASH1L knockout in PC-3M cells — lentiviral CRISPR knockout used in a bone-metastasis model.
• PTX3/TLR4 signalling in SHH medulloblastoma — lentiviral editing in DAOY cells.
• Paxillin regulation in granulosa cells — non-viral knockout in KGN cells.
• PRODH/POX in metformin-induced apoptosis — non-viral knockout in C32 melanoma cells.

• CRISPR knockout handbook
• Confirm that the sgRNA PAM requirement matches the selected Cas nuclease.
• Do not add unnecessary markers or cassettes without checking packaging limits.
• Plan genomic and protein-level validation before beginning the experiment.

Cas9-only

• Cas9, Cas9 nickase, and dCas9 vectors or viruses

sgRNA-only components

• Lentiviral sgRNA + Neomycin^R
• spCas9-compatible AAV sgRNAsp + GFP or saCas9-compatible AAV sgRNAsa + GFP
• Non-viral sgRNA + GFP
• Custom dual or multiplex sgRNA vectors

• UCP2 and muscular ATP depletion — separate AAV-Cas9 and AAV-Ucp2-sgRNA delivery in mice.
• Genome-wide CRISPR screening in HepG2 cells — stable Cas9 cells transduced with an LDLR-targeting sgRNA lentivirus.

• Avoid using the same selectable marker for both vectors unless your enrichment plan accounts for it.
• Confirm nuclease compatibility before ordering sgRNAs; spCas9 and saCas9 recognize different PAM sequences.
• Measure or enrich for co-delivery when both vectors are introduced at the same time.

• pLenti-Multi-sgRNA-PGK-Neo
• Custom dual and multiplex sgRNA vector construction

Terminology: “Dual sgRNA” usually means two guides in one construct. “Multiplex” refers more broadly to three or more guides or multiple simultaneous targets.

Large-scale dual sgRNA CRISPR library construction

abm generated more than 220 custom dual-sgRNA lentiviral vectors for a functional-genomics program. Each construct contained two sgRNA expression cassettes and a fluorescent reporter.

• 220+ sequence-verified dual-sgRNA constructs
• Consistent single-vector architecture for spCas9 workflows
• Multi-batch manufacturing and quality control
• Optional lentiviral packaging
• Suitable for pooled or arrayed screening designs

• Do not assume every guide performs equally; validate guide activity individually when possible.
• Check promoter compatibility and repeated-sequence risks when combining several sgRNA cassettes.
• For deletion experiments, design validation primers outside the intended deletion boundaries.

• spCas9: the most widely used Cas9 and a common choice for lentiviral and non-viral systems.
• saCas9: a smaller Cas9 often selected for AAV-based All-in-One designs.
• The enzymes recognize different PAM sequences, so sgRNAs are not interchangeable between them.

• Choosing a delivery method before confirming the cell type’s transfection or transduction properties.
• Exceeding viral cargo limits after adding promoters, markers, or reporters.
• Assuming AAV, lentivirus, and plasmid systems provide the same expression duration.

• Custom non-viral repair template
• All-in-One lentiviral vector with spCas9
• All-in-One AAV vector with saCas9
• All-in-One non-viral vector with spCas9

Choose the cut site and donor design together. Include silent changes when needed to reduce re-cutting after successful repair, and plan junction PCR or sequencing assays before editing.

• RFP knock-in at the human AAVS1 safe-harbour site
• Targeted integration into pseudo-attP sites in CHO cells

• Do not reuse knockout-oriented multiplex guidance for HDR donor design; the workflows have different requirements.
• Avoid placing validation primers entirely inside the donor sequence.
• Confirm whether a tag could disrupt protein localization, expression, or function.

• dCas9-VPR — transcriptional activation.
• dCas9-KRAB — transcriptional repression.
• dCas9-TET1CD — targeted DNA demethylation.
• dCas9-DNMT3A — targeted DNA methylation.
• Promoter-targeting CRISPRa/CRISPRi sgRNA vectors

• Target sgRNAs near the transcription start site or relevant regulatory region rather than a coding exon.
• Use multiple sgRNAs when practical because regulatory effects can vary strongly by target position.
• Measure the effect at the RNA level and, where relevant, the protein level.
• CRISPRi represses transcription but does not create a permanent gene knockout.

• Genetic interactions between ANLN and KDR are prognostic for breast cancer survival

• CRISPRa and CRISPRi overview
• Do not use coding-region knockout sgRNAs as a default for CRISPRa or CRISPRi.
• Confirm that the chosen sgRNA vector is compatible with the selected dCas9 effector system.