Zebrafish Floxed Gene Services

CRISPR/Cas9 is used to insert loxP sites flanking the region to be deleted in zebrafish. Insertion of two loxP sites creates the ability to permanently remove the sequences of interest with expression of the Cre recombinase which can be expressed using tissue or stage-specific promoters for precise control of gene function.
This line can then be crossed with a Cre-line expressing the Cre recombinase under a specific promoter or injected with a plasmid containing Cre recombinase. The Cre recombinase promotes recombination of the two loxP sites and the region between the sites is removed from the genome.
We can create a floxed allele for tissue-specific deletion or temporal-control of your gene of interest. This can be very useful when studying embryonic lethal genes or for understanding how your gene of interest functions in different tissues.
Conditional Mutagenesis by loxP Insertion
One of the more difficult aspects of biology is how to perform loss-of-function studies of essential genes. Now the conditional mutagenesis by loxP insertion in zebrafish is possible.
We use sequential integration of loxP sequences (e.g. two stages of injection and screening) to create "floxed" genes for large deletions of sequence in the targeted loci.
Conditional alleles
- Used to make deletion of genes in cell type and time specific manner
- Flank gene of interest with loxP sites
- Inject a plasmid or cross into strain that expresses Cre recombinase where/when/how you want loss-of-function to occur
Conditional mutagenesis by loxP insertion in zebrafish – outfoxing the unfloxable
By Ben Jussila
Floxed Gene Service Options (price reflects academic pricing)
Standard Injection Mix | Standard Injection Mix + Screening Primers | Validated sgRNA Injection Mix | Complete Validated Injection Mix | Full Build* (Sequence verified heterozygous line) | |
Floxed Gene Design | ![]() | ![]() | ![]() | ![]() | ![]() |
Injection Mix Assembly | ![]() | ![]() | ![]() | ![]() | ![]() |
Locus Evaluation & Screening Reagents | ![]() | ![]() | ![]() | ![]() |
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InVivo sgRNA Testing | ![]() | ![]() | ![]() |
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InVivo Editing Assessment | ![]() | ![]() |
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Expertly Injected Embryos | ![]() |
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Germline Transmission Screening & Line Propagation | ![]() |
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Pricing* | $1,995 | $2,995 | $4,995 | $7,995 | |
Timeline | 4-6 weeks | 4-8 weeks | 1-2 months | 4-6 months | n/a |
More Details | More Details | More Details | More Details | More Details |
Injection Mix and sgRNA Validation Details
Standard Injection Mix
Choose this option if you want us to pick your sgRNA site, have already tested your sgRNA in vivo, or want to make a CRISPR knockout. We send a custom designed injection mix and you develop your own preferred detection/screening protocol.
What’s Included?
- Consultation with a genetic engineer to develop the genome editing strategy
- In silico design of all reagents including the sgRNA site and donor homology template
- Sequence files for the donor homology construct and edited locus
- 4 vials of 10uL lyophilized sgRNAs duplexed with the Cas9 protein
- Injection dyes for early visualization of injection success
- Detailed Instructions on mix reconstitution and injection protocols.
Standard Injection Mix + Screening Primers
Choose this option if you want to inject and screen for your edit by PCR without having to worry about designing your own screening primers. This option includes our standard mix plus ready-to-go screening primers.
What’s Included?
- Our Standard Injection Mix PLUS:
- Prescreened Allele Specific PCR primers to detect your edit in zebrafish tissue.
Validated sgRNA Injection Mix
Choose this option if you want us to guarantee your injection mix is designed around a high efficiency sgRNA site using our rigorous in-house testing process.
What’s Included?
- Our Standard Injection Mix PLUS:
- Selection and testing of two sgRNAs for in vivo evaluation of cutting efficiency
- Donor homology designed around a demonstrated high efficiency sgRNA site
- In vivo screening for potential polymorphisms in your target locus
Complete Validated Injection Mix
Choose this option if you want a CRISPR knockin with completely validated reagents to use in your lab and to find your gene edited line. This is the best Custom Injection Mix option for the novice zebrafish line builder.
What’s Included?
- OUR VALIDATED sgRNA INJECTION MIX PLUS:
- Full in vivo evaluation of somatic editing efficiency.
- Confirmation of integration capacity of your edit into the zebrafish genome
Full Build (Sequence verified heterozygous line)
Choose this option if you want us to deliver a stable sequence validated CRISPR knockin line.
What’s Included?
- Veried Clutch (F0 injected Embryos) Plus:
- Growth of F0 injected embryos in our nursery
- Rigous screening of F0 injected adults for germline transmission of your edit
- Sequence conformation and propagation of F1 heterozygous animals
Injection Mix Service Component Descriptions
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