Custom cell line gene modification; KI; Timeline: 3-5 months
Cat# ASC-7102
Size : 1service
Brand : Applied StemCell
Contact local distributor :
Phone : +1 850 650 7790
Automated High-throughput Protocols for the Genetic Modification of Your iPSC Line of Choice
We can engineer your control and disease iPSC lines or choose from one of our well-characterized master iPSC lines derived from fibroblasts (male: ASE-9211; female: ASE-9209), cord blood, or PBMCs, which have proven CRISPR gene editing and differentiation potential.
| Faster Timelines with Automated High-throughput Protocols | |||
| Project type | Conventional Protocols | ASC’s Optimized High-throughput Protocols | Improvement in Delivery Times |
| Knockout (KO) | 12-20 weeks | 6-8 weeks | 60% |
| Point Mutation (Single Nucleotide Polymorphism or Variant) | 12-20 weeks | 6-8 weeks | 60% |
| Knock-in (Reporters/ Tags, Large Transgenes) | 12-24 weeks | 10-15 weeks | 20-30% |
A Variety of Other Modifications: Standard & Complex
Correct/engineer mutations or introduce a variety of genetic modifications in iPSCs:
- Gene knockout: gene disruption or site-specific large fragment knockout (>10kb)
- Gene insertion: reporter gene/ tag insertion, small fragment insertions, SNV/ point mutations
- Inducible gene expression/ gene overexpression models
- Gene fusion (translocation, inversion, etc)
Don't limit yourself to only the standard modifications. Ask us about: Multiplexed genome editing; conditional knock-in, gene fusion, and other models that you would like for your projects.
And…. We offer Customized Deliverables
- Choice of heterozygous or homozygous mutations
- Footprint-free genome editing – Ex. Single nucleotide variant (SNV; point mutation) engineering without silent mutations for regulatory compliance
- Specific genetic or safe harbor locus
CRISPR Knock-In Projects
Project 1:
Goal: Knock-in of 1 bp at the AAVS1 locus using the ASE-9211 Master iSPC Line by CRISPR/Cas9 technology
Knock-In Strategy for AAVS1 (1bp insertion)
Figure 1: Knock-in strategy for 1bp insertion in the AAVS1 locus of the ASE-9211 Master Cell Line.
Genotyping Clone #6
Figure 2: Sequencing chromatogram of iPSC line with 1bp insertion in the AAVS1 locus (top: Clone #6) compared to the Parent line, ASE-9211 (bottom).
Project 2:
Goal: Knock-in of 150bp at the AAVS1 locus using the ASE-9211 Master iPSC Line by CRISPR/Cas9 technology
Knock-In Strategy for AAVS1 (150bp insertion)
Figure 3: Knock-in strategy for 150bp insertion at the AAVS1 locus of the Master iPSC Line.
Genotyping Positive Clone #21
Figure 4: Sequencing chromatogram showing the ~150bp insertion at AAVS1 locus.
CRISPR Knockout Projects
Project 3:
Goal: 1bp deletion in the AAVS1 locus using the ASE-9211 Master Cell Line by CRISPR/Cas9 technology
Figure 5. Sequence chromatogram of iPSC line with 1 bp deletion (AAVS1-1bp DEL; bottom) compared to wild type (WT; top).
Figure 6. Sequence alignment between the 1 bp deletion iPSC line (AAVS1-1bp DEL; bottom) and wild type (WT; top).
Project 4:
Goal: 1000bp Deletion in the AAVS1 locus using the ASE-9211 Master Cell Line by CRISPR/Cas9 technology
Figure 7. AAVS1 wild type (WT) sequence showing gRNA cut sites and position of 1007 bp (~1000 bp) deletion (sequence in red).
Figure 8. AAVS1 WT chromatogram showing sites of ~1000 bp deletion (sequence in red). Top: Sequence for 5’ deletion site; Bottom: Sequence for 3’ deletion site.
Figure 9. Sequence chromatogram of iPSC line with ~1000 bp deletion in the AAVS1 locus.
Only a few NIST projects are listed, if you would like to learn more, contact us today.
- Ilic, D. (2019). Latest developments in the field of stem cell research and regenerative medicine compiled from publicly available information and press releases from nonacademic institutions in October 2018. Regenerative medicine, 14(2), 85-92.
- Simkin, D., Searl, T. J., Piyevsky, B. N., Forrest, M., Williams, L. A., Joshi, V., ... & Penzes, P. (2019). Impaired M-current in KCNQ2 Encephalopathy Evokes Dyshomeostatic Modulation of Excitability. bioRxiv, 538371. https://doi.org/10.1101/538371
- Jang, Y., Choi, J., Park, N., Kang, J., Kim, M., Kim, Y., & Ju, J. H. (2019). Development of immunocompatible pluripotent stem cells via CRISPR-based human leukocyte antigen engineering. Experimental & Molecular Medicine, 51(1), 3.
- Lizarraga, S. B., Maguire, A. M., Ma, L., van Dyck, L. I., Wu, Q., Nagda, D., ... & Cowen, M. H. (2018). Human neurons from Christianson syndrome iPSCs reveal allele-specific responses to rescue strategies. bioRxiv, 444232.
- Tanaka, H., Kondo, K., Chen, X., Homma, H., Tagawa, K., Kerever, A., ... & Fujita, K. (2018). The intellectual disability gene PQBP1 rescues Alzheimer’s disease pathology. Molecular Psychiatry, 1.
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Selvan N., George, S., Serajee, F. J., Shaw, M., Hobson, L., Kalscheuer, V. M., ... & Schwartz, C. E. (2018). O-GlcNAc transferase missense mutations linked to X-linked intellectual disability deregulate genes involved in cell fate determination and signaling. Journal of Biological Chemistry, jbc-RA118.
-
Chai, S., Wan, X., Ramirez-Navarro, A., Tesar, P. J., Kaufman, E. S., Ficker, E., ... & Deschênes, I. (2018). Physiological genomics identifies genetic modifiers of long QT syndrome type 2 severity. The Journal of clinical investigation, 128(3).
-
Seigel, G. M., et al. (2014). Comparative Analysis of ABCG2+ Stem-Like Retinoblastoma Cells and Induced Pluripotent Stem Cells as Three-Dimensional Aggregates. Investigative Ophthalmology & Visual Science, 55(13), 3068-3068.
-
Comley, J. (2016). CRISPR/Cas9 - transforming gene editing in drug discovery labs. Drug Discovery Weekly. Fall 2016; 33-48.
