Get The Correct Pup in The First Round!
How to save on you transgenic mice project? Use ssDNA as CRISPR HDR donor template for one-step, high efficiency knockin to decrease the number of mice and embryos you have to manage.
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20% OFFssDNA! Learn about ssDNA
25% OFFWhen you buy 3 or more CRISPR sgRNA or ssDNA! Promotion Details
Genetic modification of cell and animal models enables developing diagnostics strategies, identifying prognostics biomarkers, and zeroing on therapeutic targets, and CRISPR/Cas9 technology have simplified and speed up the workflow significantly. CRISPR/Cas9 induces double-strand DNA breaks at gRNA target sites and activates cellular repair processes, including the homology-directed repair (HDR) pathway which has been exploited to knock-in specific sequences with exogenously provided donor DNA template. There are a variety of HDR donor formats (plasmid, dsDNA, ssDNA) for the generation of CRISPR-edited mice, however their editing efficiency differs significantly.
HDR Donor Options:
- Plasmids DNA – Editing Efficiency : 10-30%, but requires very long homology arms1,2
- dsDNA – Editing efficiency: 1- 10%3-8
- Two ssODN for LoxP insertion – Editing Efficiency < 10%9-11
- ssDNA – Editing Efficiency ~ 30-60%6-9
"We found that insertion efficiency was 2.5 times higher, and the viability of injected embryos was 2.4 times higher using an ssDNA donor than a dsDNA donor." - Miura, H., Quadros, et al. Easi-CRISPR for creating knock-in and conditional knockout mouse models using long ssDNA donors. Nat Protoc 13, 195–215 (2018).
"lssDNA generally provides more efficient KIs of approximately 10–50% in zygotes compared with double stranded DNA that normally provides less than 10%." - Miyasaka, Y., et al. CLICK: one-step generation of conditional knockout mice. BMC Genomics 19, 318 (2018).
"We test some of the newer methods that use one-donor DNA on 18 loci for which the two-donor approach failed to produce cKO alleles. We find that the one-donor methods are 10- to 20-fold more efficient than the two-donor approach." -Gurumurthy, C.B., et al. Reproducibility of CRISPR-Cas9 methods for generation of conditional mouse alleles: a multi-center evaluation. Genome Biol 20, 171 (2019).
What does high editing efficiency mean to your projects?
- 2-3 fold increase in KI efficiency with ssDNA compared to plasmids, dsDNA, and short ssODNs.
- 2 X Editing Efficiency = 50% Less Mice, 50% Less Injection, 50% Less Sequencing.
- Save 6-8 weeks with one less breeding round using one step knockin with long ssDNA vs. two ssODNs.
In general, ssDNA have been accepted as the preferred donor formats to maximize on-target integration for conditional KO/KI and larger KI animal model generation. The increased availability of long ssDNA in recent years has certainly made these methods more feasible.
Recommended Package for Conditional KO/KI or Large Gene Insertion
EasyEdit sgRNA and Cas9 Protein- No annealing necessary
- Modified for improved stability and efficiency
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ssDNA Synthesis Service- Higher KI efficiency than other options
- Lower cytotoxicity than dsDNA
- Sequences up to 5 kb, 100% Sequence fidelity confirmed by Sanger
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Promotion Terms and Conditions:
- The promotion is only valid for our standard EasyEdit sgRNA (SC1969), SafeEdit sgRNA (SC1968) and ssDNA (SC1999) services.
- If you order 1 or 2 ssDNAs, a 20% discount will be automatically applied to your SC1999 item(s).
- You can mix and match any EasyEdit sgRNA, SafeEdit sgRNA or ssDNA in your order. If you order 3 or more sgRNA or ssDNA items, or combination thereof, a 25% discount will be automatically applied to all SC1969, SC1968, and SC1999 items.
- This promotion cannot be combined with any other promotions or special pricing agreements.
- Promotion is effective only for orders placed between Nov. 1st to Dec 31st , 2020.
- Promotion is valid for domestic (US) and international customers, excluding Japan and China.