Transcription profiling of Xenopus laevis developing notochord

We screened for differentially expressed genes in the developing notochord using the Affymetrix microarray system in Xenopus laevis. At late gastrula, we dissected four regions from the embryo, anterior mesoderm, posterior mesoderm, notochord and presomitic mesoderm. Three types of comparison were carried out to generate a list of predominantly notochord expressed genes: (1) Posterior mesoderm vs. anterior mesoderm; notochord genes are expected to be increased since the notochord is located in the posterior mesoderm. (2) Posterior mesoderm vs. whole embryos; notochord genes are expected to be increased. (3) Notochord vs. somite. This comparison sub-divided the group of posterior mesodermal genes identified in (1) and (2). All tissues are dissected using tungsten needles. We first dissected dorsal tissue above the archenteron from late gastrula to early neurula. To loosen tissue, we treated the dissected dorsal explant in a 1% cysteine solution (pH 7.4) and removed the neuroectodermal layer. Anterior mesoderm was dissected corresponding to about the anterior one-third of the archenteron roof, and the rest was collected as posterior mesoderm. The posterior mesodermal explant was dissected into notochord and somites, following a clearly visible border between the two tissues. The accuracy of all dissection was confirmed by RT-PCR of marker genes.

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Tanegashima K, Zhao H, Rebbert ML, Dawid IB