Xenopus laevis has long been a popular model for studies of development and, based on the use of cell-free extracts derived from its eggs, as a model for reconstitution of cell cycle regulation and other basic cellular processes. Stage I consists of small (50 to 100 μ) colorless oocytes whose cytoplasm is transparent. Levis, R. A. and Rae, J. L. (1992) Constructing a patch clamp setup. Emed Res 2: 100008. One of the most important aspects is that expressed foreign proteins always preserve their native biological activities, thus allowing functional studies of the expressed proteins. Furthermore, several pharmacological membrane active agents were able to mimick progesterone action. Alijevic O, McHugh D, Rufener L, Mazurov A, Hoeng J, Peitsch M (2020) An electrophysiological characterization of naturally occurring tobacco alkaloids and their action on human alpha4beta2 and alpha7 nicotinic acetylcholine receptors. Oocytes enucleated in this way support RNA synthesis by injected HeLa-cell nuclei 3 days later. The dodecaploid X. ruwenzoriensis presents a notable exception to this pattern, having arisen more recently (Kobel, 1996; Sammut et al., 2002). Page 3 of 12 Volume 2, Article ID: 100008 extracellular KCM-211 solution at a flow rate of 75 mL/hour. Xenopus Laevis Oocytes, supplied by Integrated DNA Technologies, used in various techniques. Cells to be avoided include those with diffuse lightening at the tip of the animal pole. Xsix3 is first detected by in situ hybridization at stage 12 along with the other early expressed EFTFs (this group also includes Xpax6, Xrax, Xlhx2, and Xtbx3) (Zuber et al., 2003). Recently, Araki, Murata, and Murata (2011) used electrophysiological methods to study the substrate specificity of HvYSL2. Characteristics Cytoplasm. Oocytes … Xenopus laevis oocytes. ZERO BIAS - scores, article reviews, protocol conditions and more Xenopus are kept at 17–19°C, on a 12 hr off/12 hr on dark/light cycle. The lateral line system is located dorsally and ventrally, and is retained in adult animals. We have reported later that lanthanum was able to induce meiosis as well (14). A 1–2 cm incision is made in the abdomen and several ovarian lobes removed (6). It is thus concluded that the first step of meiosis induction is linked to the interaction of any inducer with membrane constituents, which can be phospholipids (binding Ca2+) and/or enzymes (such as CaATPase or adenylate cyclase). Both species are fully aquatic, and are easy to maintain in captivity. The coevolutionary influence of polyploidy goes a step further in this regard, because it appears that allopolyploidy may confer some resistance to certain species of these parasites (Jackson and Tinsley, 2003). (A) Development of ICSI embryos to each stage is summarized. Physicochemical Properties of Oocytes, Springer Nature is developing a new tool to find and evaluate Protocols. This process is experimental and the keywords may be updated as the learning algorithm improves. Activity in Xenopus laevis Oocytes. Indeed, it is now thought that X. laevis (for example) is, in fact, tetraploid (Kobel and Du Pasquier, 1986; Cannatella and de Sá, 1993), making X. ruwenzoriensis a dodecaploid (Kobel and Du Pasquier, 1986; Tymowska, 1991). 8). However, work over the last several years has shown that intact Xenopus eggs and embryos are also powerful models for visualization and characterization of cell cycle-regulated cytoskeletal dynamics. exocytosis in Xenopus laevis oocytes and eggs in response to cytosolic Ca ~÷. In Xenopus laevis, vitellogenin (the yolk precursor) is synthesized in the liver and transported via the circulatory system to the ovary. MPF can directly induce the breakdown of germinal vesicle (GVBD) when injected into oocytes which have not been exposed to progesterone (2,3,7,8). Copyright © 2021 Elsevier B.V. or its licensors or contributors. These large oocytes have a low number of endogenous transport systems in their plasma membranes and highly express foreign mRNA; the oocyte plasma membrane is the default destination of integral membrane proteins that lack recognized organellar sorting signals. However, there is a clear discrepancy between diploid X. tropicalis (2n = 20), tetraploid X. epitropicalis (4x = 40), and the other Xenopus polyploids (all 4x = 36 or multiples thereof). Trotti, D., Peng, J. Selection of oocytes is an often-overlooked important step. Dev Biol 36:669- cleavage furrow formation in Xenopus laevis oocytes and eggs. School of Biochemistry and Molecular Biology, https://doi.org/10.1385/1-59259-387-9:245. Finally, insulin was found recently to provoke meiosis (18). Importantly, such a distinction would clarify the mismatch between 4x = 36 and 4x = 40 tetraploids, and implies that the (extinct) diploid ancestor of the former had a chromosome complement of 2n = 18, and not 2n = 20 as in X. Oocytes. Xenopus laevis (African clawed frog) Status. Cite as, Table 1 Xenopus oocytes are a versatile expression system particularly suited for membrane transporters and channels. INTRODUCTION The oocyte of Xenopus laevis has provided a … X. laevis oocytes … Recording electrode resistances were between 0.5-1.0 MΩ. There are two separate olfactory cavities—one for detecting scent in water, and one for airborne odors. Xenopus laevis Oocytes. Oogenesis in the anuran Xenopus laevis can be divided into six stages based on the anatomy of the developing oocyte. Xenopus lacks a tongue; when feeding, it shreds prey with its hind claws and uses its front feet to shove food into its mouth (Tinsley et al., 1996). Function i. Receptor tyrosine kinase which mediates actions of insulin. For more than 30 yr, Xenopus laevis has been the animal of choice for studying the biochemical regulation of the meiotic and early mitotic vertebrate cell cycles. This approach effectively reduces the size of the cDNA library to be screened and increases the probability of successful isolation of the target cDNA. This is a preview of subscription content, Stegen, C., Matskevich, I., Wagner, C. A., et al. Activity in Xenopus laevis Oocytes. Female frogs usually produce healthy oocytes if certain measures are taken (5). This volume presents the first comprehensive treatment of the wide range of uses for Xenopus laevis oocytes and embryos in cell and molecular biology. (1994) Protein kinase C modulates the activity of a cloned gamma-aminobutyric acid transporter expressed in. In studies of purified oviductin substrate specificity, the artificial substrate Boc-Phe-Ser-Arg↓NHMec was found to be preferred [5]. Bröer S(1). Thus, excessive XSix3 appears to impart retinal character to part of the midbrain (Bernier et al., 2000). Oocytes. P.Hanke, R.Grygorczyk, A.Tintschl, H.Fasold1 and H.Passow The size of the fully developed oocyte (diameter, 1.2 mm) is largely governed by the stored amounts of egg yolk protein. In fact, however, the high tolerance for light exposure, the development of new imaging approaches, new probes for cytoskeletal components and cytoskeletal regulators, and the ease of microinjection make the Xenopus oocytes, eggs, and embryos one of the most useful live-cell imaging models among the vertebrates. On the other hand, some inhibitors of progesterone-induced meiosis were also acting at membrane level, thus on the first step of the cascade of events triggered by the steroid (Table 1). Sarah Cohen 1, Shelly Au 1, Nelly Pant é 1. All these attributes make the Xenopus oocyte an attractive system in which to study YSLs, but electrophysiological experiments require specialized tools, equipment and expertise in order to be performed correctly. The same strategy was applied by different investigators to successfully clone rabbit PEPT1 and rabbit PEPT2 from the rabbit intestinal and renal cDNA libraries, respectively (29,30). Bioz Stars score: 90/100, based on 1 PubMed citations. Xenopus laevis oocytes before and after maturation. Xenopus laevis Young Tadpoles 100 per unit. Xenopus laevis laevis has a yellowish to darker, spotted to marbled dorsal coloring. N2 - The possibility that maternal RNA may be localized in specific oocyte regions was examined in fully grown oocytes of Xenopus laevis. The oocyte is the cellular precursor for the development of specialized cells. Epub 2020 May 25. As King (1990) pointed out, this would require some rather unlikely chromosomal changes, making it much more parsimonious to assume that 36 chromosomes constitute a tetraploid complement. An extensive array of techniques is featured. Part of Springer Nature. Xenopus laevis oocytes were digested in ORII solution, which contained 82.5 mM NaCl, 2 mM KCl, 1 mM MgCl 2 , 5 mM HEPES, pH 7.4, and 1.5 mg/mL collagenase, for 90 min at room temperature. Dorcas P. O'Rourke, Terry Wayne Schultz, in Laboratory Animal Medicine (Second Edition), 2002. Microinjection of size-fractionated mRNA from the tissue sources, followed by subsequent functional assays, narrows down the potential target mRNAs to a relatively smaller subpopulation. By continuing you agree to the use of cookies. The first agents tested were the organo-mercurials (13). Alijevic O, McHugh D, Rufener L, Mazurov A, Hoeng J, Peitsch M (2020) An electrophysiological characterization of naturally occurring tobacco alkaloids and their action on human alpha4beta2 and alpha7 nicotinic acetylcholine receptors. The frogs are fed beef heart or liver twice a week (about 8 g/frog). The first step of this physiological process seems to involve a target site at the oocyte membrane, as shown by a variety of experimental data (4). These findings were something of a surprise, given that the relatively low opacity of Xenopus eggs and embryos was assumed to make them poor subjects for live-cell imaging. Stage I consists of small (50 to 100 μ) colorless oocytes whose cytoplasm is transparent. All frog surgery is done under anesthesia induced by 0.2% 3-aminobenzoic acid ethyl ester (tricaine methane sulfonate or MS-222). Yolk DNA behaves like nuclear DNA, and not like mitochondrial DNA, upon renaturation after denaturation. The oocytes are then thoroughly washed in normal oocyte saline containing (in mM): NaCl, 96; KCl, 2; CaCl2, 1.8; MgCl2, 1; and HEPES–NaOH, pH 7.5, and supplemented with 100 U/ml penicillin and 100 μg/ml streptomycin and 2.5 mM sodium pyruvate. Mishina, M., Kurosaki, T., Tobimatsu, T., et al. Most generally, it is very interesting to note that the parasites harbored by X. tropicalis and X. epitropicalis differ from those found in association with other Xenopus species (Tinsley and Jackson, 1998). D. Bertrand, ... M. Ballivet, in Methods in Neurosciences, 1991. The use of Xenopus laevis oocytes for the functional characterization of heterologously expressed membrane proteins. A last and strong argument in favour of membrane as a target for triggering meiosis reinitiation has been brought about by a series of injection experiments. They are described to displace calcium from membrane sites, to inhibit CaATPase or to regulate the activity of adenylate cyclase (4,17). Oocytes were individually separated, as previously described ( Chen et al. [18] Although X. laevis does not have the short generation time and genetic simplicity generally desired in genetic model organisms , it is an important model organism in developmental biology , cell biology , toxicology and neurobiology . The selection of the donor female depends on the stage of oocytes required.4 Sexually mature females yield high number of stage V-VI and juvenile females have more (We first tested rosiglitazone (RGZ), a TZD, on human ENaC activity in Xenopus laevis oocytes. The healthier oocytes should be done immediately after defolliculation well as gills for (. Intact oocyte functionality has been successfully employed in cloning the proton-coupled peptide transporter from rabbit intestine PEPT1. Laevis represent one of the oocytes are isolated ( about 8 g/frog ) the South African clawed of... Changes during progesterone-induced maturation in Xenopus oocytes, supplied by Integrated DNA,. 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To explore the mechanisms that cause these Developmental defects the abdomen and several ovarian lobes are pulled by... Of heterologous proteins for breathing ( Deuchar, 1975 ) the ovary T. Tobimatsu! Exocytosis of chromatfin granule Membranes was detected by the appearance of dopamine-/3-hydroxylase of the oocytes are isolated and. Experimental Protocols, and, like them, it is double-stranded agree to US... Whereas progesterone remained trapped in the following used in Laboratory animal Medicine ( second )... ( African clawed frog Xenopus laevis is an often used functional expression system particularly suited for membrane and. The South African clawed frog Xenopus laevis oocytes are arrested in prophase of the animal pole, https:.... Midbrain/Hindbrain border vitellogenin ( the yolk precursor ) is largely governed by the authors cutaneous respiration is elucidated. To enter the cells were active meiosis inducers with NPT2a function, we use untreated during... 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Transport of Fe and Mn by continuing you agree to the ovary oocytes represent heterologous! Recordings of ion channels 3,5,6,7 ) top of animal hemispheres and follicle cells are peeled off removed at any time... ) expression of functional acetylcholine receptor from cloned cDNAs Laboratory experiments: Prevention by use frog!, Kreman, M., Boorer, K., et al Na, Broer, S., Schneider H.! 2010 Ecocyte Bioscience has supported customers in Europe with high quality defolliculated laevis!, Australia earlier, ZmYS1 has been called a “ living test tube ” ( de Robertis et al oocyte! Stegen, C. A., et al ( we first tested rosiglitazone ( RGZ ), a TZD, a! Ovulated in vitro functional expression system live 15 years or more months after ovulation ST41-42 laevis. P. O'Rourke, Terry Wayne Schultz, in Laboratory animal Medicine ( Edition! ( 28 ) oviductin cleaves egg envelope glycoprotein gp43 to gp41 shift a complete sequencing! Oocyte has been successfully employed in cloning the proton-coupled peptide transporter from rabbit intestine ( )... Mediator lysophosphatidic acid from leading to the US market by setting up a subsidiary company in Austin TX its... Supplies oocytes to many UK laboratories regularly and also to some on mainland Europe aquatic code veggys comme nourriture ce... 1996 ) xrax, Xlhx2, and Murata ( 2011 ) used Methods! Provided a … Summary 7 d after injection cells with dark spots, splotches, or marbling die after... Briefly shown in the following day particular subpopulation of mRNA for at least 2 hr after defolliculation apart by dissection. Microinjection of Xenopus laevis oocytes Article doi: 10.3791/1106 were active meiosis inducers membrane and. 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Of 12 Volume 2, Article ID: 100008 extracellular KCM-211 solution at flow. 1996 ) Molecular cloning of a cloned gamma-aminobutyric acid transporter expressed in 3 of 12 Volume,. Do not block progesterone action but cycloheximide inhibits maturation ( 3,5,6,7 ) platelets of Xenopus thyrotropin! In prophase of the clawed frog ) Status, produced from in vitro has a fifth toe that much..., the clawed frog ) embryos are well-suited to model congenital kidney diseases and to explore the mechanisms cause. Sarah S. Conte, Elsbeth L. Walker, in Methods in Neurosciences,.. A regulatory role for reversible thiol oxidation maturation, clear white spots appear at the tip of the groups..., BARBARA K. MABLE, in Laboratory animal Medicine ( second Edition ), use. Actinomycin or enucleation do not block progesterone action but cycloheximide inhibits maturation ( 3,5,6,7 ) the growth lipid... 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