The nematode cuticle is the animal's external skeleton and plays important roles in the maintenance of body morphology and integrity, protecting the animal from the external environment and animal's motility. During the development of the nematode, the cuticle is synthesized five times in total, including the first time in the embryo and the other four times at the end of larval stages. The cuticle is a highly complex structured extracellular matrix (ECM), which predominantly consists of cross-linked collagens, additional insoluble proteins (cuticlins) and relevant glycoproteins and lipids. As an absolute proportion of cuticle, the collagens are widely studied, involving its associated genes and the process of biosynthesis. In addition, as a highly specialized ECM, the nematode cuticle has distinct and complex features. Even so, the biogenesis of cuticles with respect to molecules and pathways is still conserved in vertebrates. Here, we provide C. elegans cuticle isolation service using two protocols referenced in previous studies to offer customers further insight into C. elegans' cuticle collagen content and other related research topics.
The nematode Caenorhabditis elegans is enclosed within the cuticle in post-embryonic development. The cuticle is a collagenous ECM synthesized by the hypodermis (an underlying ectodermal cell layer) that surrounds the body of the nematode. The cuticle surrounding the main body of the nematode can be divided into the dorsal and ventral regions and further exhibits a gross multilayered ultrastructure that varies in detail at different developmental stages. Besides, there are narrow lateral regions overlying the seam cells where longitudinal ridges are termed as alae, which are distinct from protein composition and ultrastructure of the cuticle. Moreover, the major layers of the cuticle include the outer epicuticle layer, cortical, medial and inner basal layer. There are circumferential ridges (annuli) that exist on the cortical layer of the dorsal and ventral regions.
The organization and structure of the C. elegans cuticle. (Page, A.P., et al, 2007)
The major component of the C. elegans cuticle is collagen, a structural protein that is ubiquitous. The cuticle collagens have characteristic [Gly-X-Y] repeats, which are encoded by a gene family with over 170 members. At present, mutants of 21 of the cuticle collagen genes have been identified, involving a number of informative body morphology defects. Apart from collagens, a highly cross-linked insoluble class of proteins termed cuticlins are implicated with the cuticle. Moreover, the outermost layer, the epicuticle is lipid-rich and covered by the glycoprotein-rich negatively charged surface coat.
The C. elegans cuticle can be isolated largely intact and free of all cellular material through treatment with 1% sodium dodecyl sulfate (SDS) and sonication. Here we offer two modified methods for customers to isolate the C. elegans cuticle.
The freeze-thaw protocol is relatively simple. In addition to its simplicity, the SDS-cleaned cuticles are suitable for further desired analysis. It is notable that before starting the cuticle isolation, C. elegans are age synchronized and hypochlorite treatment is adopted. Here, in order to provide better service, we adopt glass materials instead of plastic materials to avoid C. elegans cuticle sticking to the plastic tube during the freeze-thaw procedure.
This method is designed for obtaining pure cuticles from synchronous populations of C. elegans adults. The ST buffer is composed of 1% SDS, 0.125 M Tris-HCl and ddH2O, and the final PH is adjusted to 6.8. In order to offer high-quality experimental service, we conduct the procedure strictly. For instance, the steps after the sonication should be carried out on ice, except steps involving SDS.
Creative Biogene is a dedicated provider of C. elegans model services. The extreme genetic tractability makes C. elegans an excellent experimental model system for the dissection of ECM formation. Here we provide the two C. elegans cuticle isolation methods and the services are tailored to meet customers' specific needs. Creative Biogene is a reliable partner dedicated to accelerating the speed to achieve customers' research goals. Please feel free to contact us for more information about C. elegans cuticle isolation service.
* For research use only.