C. elegans Knock-in Services
The nematode Caenorhabditis elegans is widely used in genetic and biomedical research. DNA fragment insertions can be introduced into the C. elegans genome. Our C. elegans knock-in services include point mutations, floxed alleles, degron tagging, fluorescent tagging and immunotagging.
In the past 5 years, InVivo Biosystems has made over 600 fluorescent transgenic lines includuing 15 different fluorophores, 8 of which are variants of GFP.
Fluorescent Tagging. Intestinal nuclei from late L4 worms in an lmn-1p::GFP::LMN-1 transgen
C. elegans Knock-in Service Offerings
Fluorescent Tag
Visualize your protein via addition of a fluorescent protein tag at the native locus.
Immunotag
Use your protein for biochemical studies by adding an immunotag at the native locus.
Point Mutation
The best gene editing method for creating small, precise edits to introduce a small number of nucleotide changes at a target site.
Whole Gene Humanization
Replace a C. elegans gene with an orthologous gene from human or any other organism.
C. elegans Knock-in Service Pricing
| Build Type | Full Build | Candidate Lines | Custom Injection Mix |
| Transgenic Design |  | | |
| Injections | | | |
| Candidate Screening | | | |
| Strain Confirmation | | ||
| Pricing | $3,465 and up | $2,435 and up | $1,740 and up |
| Timeline | 8 weeks and up | 6 weeks and up | 4 weeks and up |
C. elegans Knock-in Service Details
C. elegans Point Mutation Service
Our Point Mutation service uses CRISPR/Cas9, which is the best gene editing method for creating small, precise edits to introduce a small number of nucleotide changes at a target site.
With this service you can:
- study a disease-causing mutation
- humanize a critical amino acid
- explore the binding site of an enzyme
- introduce phosphomimetics
- mutate isoform start sites or make any specific mutation of interest.
Our CRISPR/Cas9 Editing Is Typically Very Efficient
Among the F1 candidates with the Co-CRISPR edit, our average percentage of animals with the target edit was 65.7%.
Edit percentages as high as 95% were observed.
We Can Generate Point Mutations Quickly
For these projects, our average time from injection to sequence confirmation of two independent homozygous lines was 15.3 days.
Our quickest project only took 10 days!
96 Point Mutations In 1 Gene
All 96 mutations were created in the STXBP1 gene (which is associated with epilepsy in humans) via CRISPR.
The worm homolog of STXBP1, unc-18, causes uncoordination and near-complete lack of pharyngeal pumping when knocked out.
The functionality is restored by replacing the worm gene with the coding sequence for human STXBP1.
C. elegans Fluorescent Tagging Service
Fluorescent tagging of genes is widely used in many model organisms to study which tissues a gene of interest is expressed in, where in the cell a gene is expressed, or to determine whether multiple genes of interest are expressed in the same location. C. elegans is particularly well suited to fluorescence studies because they are transparent. Fluorescence can be easily observed in both live and fixed images.
Knock-in a fluorescent protein at the endogenous locus using CRISPR gene editing technology.
Adding a fluorescent protein tag at the endogenous locus enables you to:
- detect your protein in-vivo at native gene expression levels
- ensure that you have the correct expression pattern
- identify protein-protein interactions
- confirm degradation of your protein when paired with a degron sequence
- visualize protein synthesis rates
- quantify protein levels
- visualize protein localization to organelles or membranes
Choose From a Variety of Fluorophores
You can choose from a variety of fluorophores including YFP, BFP, eGFP, mCherry, mOrange, mScarlet, and many more. All of our fluorescent fluorophores are optimized for expression in C. elegans via codon optimization and insertion of introns.
Use our fluorescent fluorophore tags in combination with the Fluorescent Protein RNAi to reduce or eliminate fluorophore expression.
Concerned that the addition of a fluorescent fluorophores will inhibit protein function? We can use a 2A or SL2 sequence to separate the 2 components!
Things to consider when ordering your strain:
- Fluorescent protein best suited for your experiments
- Best terminus to tag
We are not limited to a fixed list of fluorophores. If you have a fluor that you would like to use, you can provide us the sequence and we will incorporate it into the design of your transgenic project. If you need your fluorescent transgenics lines to be imaged, we can readily help you with that too! If you are unsure, our genetic engineering experts will be glad to advise you. Contact us.
For immobilization of live animals to image yourself, learn more about NemaGel.
C. elegans Immunotag Service
The addition of an immunotag at the endogenous locus enables you to quantify your protein without altering the level of gene expression.
Immunotags are added using CRISPR/Cas9 gene editing technology.
You can choose from a variety of tags including FLAG, HA, HIS, TAP, or S-peptide.
All tags have been optimized for expression in C. elegans.
C. elegans Degron Tagging Service
You can knock-in a degron tag on an endogenous protein or fluorescent protein with our degron tagging service.
Adding a degron tag enables you to:
- use protein degradation as an complementary method to genetic knockouts or RNAi
- tag your protein of interested for degradation at the protein level
- control the timing of protein degradation with an inducer (e.g. auxin, blue light)
- confirm degradation of your protein when paired with a fluorescent sequence
Choose From a Variety of Systems
You can choose from a variety of degron inducible and developmental systems including Auxin Inducible Degradation, Photosensitive or blue-light inducible degradation, degron-tagged reporters of membrane topology, and OMA-1 tags that work to degrade protein from the 1-cell stage.
Use our degron tags in combination with the fluorescent tagging to visualize protein expression and degradation.
Things to consider when ordering your strain:
- Degron tag best suited for your experiments
- Best terminus to tag
For auxin-inducible degradation, TIR1 is necessary to generate a functional recognition complex and achieve successful target degradation. If you do not have a TIR1-expressing strain ready, we can suggest the most suitable commercially available strain, or we can generate a custom TIR1 strain for you.
If you are unsure, our genetic engineering experts will be glad to advise you. Contact us.
C. elegans Whole Gene Humanization
Whole gene humanization allows you to replace a C. elegans gene with an orthologous gene from human or any other organism. If the human gene rescues the function of the gene deleted-KO, you know there is conserved biology. This is often the first step in a project to create a model to study clinical variants.
We take care to design our humanization projects to preserve endogenous transcription signals so the expression patterns and levels will be maintained. In addition, we have a proprietary sequence optimization protocol that we use to create a transcript that will be expressed well in C. elegans.
Knock-in your human (or other animal) gene of interest at the locus of an orthologous gene using CRISPR gene editing technology.
Creating a whole gene humanized line enables you to:
- Determine if the human protein is functionally equivalent to the C. elegans protein.
- Study drug effects on the human protein in an animal model.
- Create a model for the study of clinical variants.
C. elegans Whole Gene Humanization Service Pricing
| Service Package | Price | Est. Delivery Time |
|---|---|---|
| Full Build | $5,105 and up | 8 - 12 Weeks |
| Candidate Lines | $4,024 and up | 6 - 8 Weeks |
| Custom Injection Mix | $2,295 and up | 4 - 6 Weeks |
Resources
Publications
Point Mutation
Repurposing the aldose reductase inhibitor and diabetic neuropathy drug epalrestat for the congenital disorder of glycosylation PMM2-CDG
Sangeetha Iyer, Feba S. Sam, Nina DiPrimio, Graeme Preston, Jan Verheijen, Kausalya Murthy, Zachary Parton, Hillary Tsang, Jessica Lao, Eva Morava2 and Ethan O. Perlstein. Disease Models & Mechanisms (2019) 12.
Tagging
Disruption of the Caenorhabditis elegans Integrator complex triggers a non-conventional transcriptional mechanism beyond snRNA genes
Gómez-Orte E, Sáenz-Narciso B, Zheleva A, Ezcurra B, de Toro M, López R, Gastaca I, Nilsen H, Sacristán MP, Schnabel R, Cabello J. PLoS Genetics. 2019 Feb 26;15(2):e1007981.
Olfactory stem cells reveal MOCOS as a new player in autism spectrum disorders
Féron, F; Gepner, B; Lacassagne, E; Stephan, D; Mesnage, B; Blanchard, MP; Boulanger, N; Tardif, C; Devèze, A; Rousseau, S; Suzuki, K; Izpisua Belmonte, JC; Khrestchatisky, M; Nivet, E; Erard-Garcia, M. Mol Psychiatry. 2016 Sep;21(9):1215-24.
Regulation of DJ-1 by Glutaredoxin 1 in Vivo: Implications for Parkinson’s Disease
Johnson, WM; Golczak, M; Choe, K; Curran, PL; Miller, OG; Yao, C; Wang, W; Lin, J; Milkovic, NM; Ray, A; Ravindranath, V; Zhu, X; Wilson, MA; Wilson-Delfosse, AL; Chen, SG; Mieyal, JJ. Biochemistry. 2016 Aug 16;55(32):4519-32.
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