Microarray-based analysis of early development in Xenopus laevis.
In order to examine transcriptional regulation globally, during early vertebrate embryonic development, we have prepared Xenopus laevis cDNA microarrays. These prototype embryonic arrays contain 864 sequenced gastrula cDNA. In order to analyze and store array data, a microarray analysis pipeline was developed and integrated with sequence analysis and annotation tools. In three independent experimental settings, we demonstrate the power of these global approaches and provide optimized protocols for their application to molecular embryology. In the first set, by comparing maternal versus zygotic transcription, we document groups of genes that are temporally regulated. This analytical approach resulted in the discovery of novel temporally regulated genes. In the second, we examine changes in gene expression spatially during development by comparing dorsal and ventral mesoderm dissected from early gastrula embryos. We have discovered novel genes with spatial enrichment from these experiments. Finally, we use the prototype microarray to examine transcriptional responses from embryonic explants treated with activin. We selected genes (two of which are novel) regulated by activin for further characterization. All results obtained by the arrays were independently tested by RT-PCR or by in situ hybridization to provide a direct assessment of the accuracy and reproducibility of these approaches in the context of molecular embryology.
Altmann
C R
CR
Laboratory of Molecular Vertebrate Embryology, The Rockefeller University, 1230 York Avenue, New York, New York 10021, USA.
Bell
E
E
Sczyrba
A
A
Pun
J
J
Bekiranov
S
S
Gaasterland
T
T
Brivanlou
A H
AH
D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D015536:Down-Regulation; D005821:Genetic Techniques; D017403:In Situ Hybridization; D008962:Models, Theoretical; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D015854:Up-Regulation; D014981:Xenopus / Q000196:embryology*
D018076:DNA, Complementary