TARGATT™ Site-Specific Knock-in Cell Line Service
TARGATT™ Fast and Site-Specific Gene Insertion in Mammalian Cell Lines
Do you need a precise comparison of different genes from more than one cell line? Do you want to generate a master cell line expressing different reporter genes? TARGATT™ provides transgene expression and a knock-in cell line in one step.
Generation of TARGATT™ master knock-in cell line with attP docking sites allows:
Site-specific integration at pre-defined loci.
High integration efficiency with no disruption of endogenous genes
Single-copy integration and stable expression
Single-step transfection to create stable cell lines
No massive clone screening required
Inclusion of large transgenes up to 20 kb
Applied StemCell is now providing a TARGATT™ master knock-in cell line generation service
You send us your favorite cell lines
We will insert the attP docking sites at specific loci
Fast: get your transgenic cell line in 3 months!
Need Site-Specific Knock-in of iPSCs? Applied StemCell can do that too!
Site-Specific Knock-in of TARGATT™ sites
Figure: Using mCherry reporter (left) in human iPSC line. Original Cell Line: hiPSC (right).
FAQ for Gene Knock-in Technology and TARGATT™ Master Knock-in Cell Line Service
1. I am interested in making a master TARGATT™ cell line using my own cell line as a parental cell line. Can you take my cancer cell line, for example, to generate a master cell line using your service?
2. What is the Gene Knock-in Technology used to generate master cell lines?
We use CRISPR/Cas9 to generate master cell lines by inserting TARGATT™ attP sites at a desired locus.
3. How long does it take to make a master cell line?
3-5 months, depending on the nature of the cell line.
4. Is there a size limit on DNA to be inserted into the genome (to attP site)?
So far we have successfully inserted a transgene of 20kb using this method.
5. What is the final deliverable product?
We ship at least 2 vials, each at 0.5x10^6 cells/vial, cryopreserved cells per clone with a report of the project. Additional clone(s) and vial(s) are available upon request. For fee-for service projects, you can also have the CRISPR and TARGATT™ vectors upon request.
6. What safe harbor loci are available to place the TARGATT™ docking site? Can you recommend one for my cell lines?
We have several ready-to-use safe harbor loci to choose from. They are Rosa26, H11, and AAVS1. Docking site insertion can be customized to other loci base on your project.
7. I got my master cell line. Do you provide plasmid so that we can construct Knock-in vector with our gene of interest?
8. Do you have off-the-shelf TARGATT™ Master cell lines?
Currently, we have human iPSC and mouse C57/BL6 ESC and iPSC lines. CHO cells will be launched soon. We are happy to discuss for other cell lines as Custom Cell Line Services.
Description of the technology
- Zhu, F., et al. (2014). Nucleic acids research, 42(5), e34-e34.
- Tasic, B., et al. (2011). PNAS USA, 108(19), 7902–7907.
Commentary, comparison with other transgenic methods
- Rossant, J., et al. (2011). PNAS USA 108(19), 7659–7660.
Tet inducible mice generated by TARGATT™
- Fan, X., et al. (2012). Endocrinology, 153(11), 5637–5644.
Advantage of Hipp11 (H11) locus
- Hippenmeyer, S., et al. (2010). Neuron, 68(4), 695–709.
Application for mice generated by TARGATT™
- Booze, M. L., et al. (2016). Free Radical Biology and Medicine, 99: 533-543.
- Feng, D., et al. (2016). The Journal of Clinical Investigation, 126(6).
- Park, K. E., et al. (2016). International journal of molecular sciences,17(6), 810.
- Sun, N., et al. (2015). Measuring in vivo mitophagy. Molecular cell, 60(4), 685-696
- Guenther, CA., et al. (2014). Nature genetics, 46(7), 748-752
- Devine, WP., et al. (2014). eLife, 3, e03848.
- Villamizar, C. A. (2014) UT GSBS Dissertations and These (Open Access). Paper 508 (2014)
- Fogg, PC., et al. (2014). Journal of molecular biology, 426(15), 2703-2716