Molecular Reproduction and Development · 2004 · 39 citations · 28 references
Life SciencesDevelopmental BiologyMolecular BiologyEmbryologyGene ExpressionGenomicsAnimal GenomicsBlastocyst DevelopmentDna SequencingMammalian EmbryologyPolymerase Chain ReactionPreimplantation Embryo DevelopmentBovine CloningDevelopmental Stage-specific ExpressionExpressed Sequence Tags AnalysisGene Expression In BlastocystsReverse Transcription Polymerase Chain Reaction (Rt-pcr)Sequence Analysis Of Cloned GenesSequence SimilarityTranscriptomics Of Blastocysts
The identification of embryo-specific genes would provide insights into early embryonic development. However, the current methods employed to identify the genes that are expressed at a specific developmental stage are labor intensive and suffer from high rates of false positives. Here we employed a new and accurate reverse transcription-polymerase chain reaction (RT-PCR) technology that involves annealing control primers (ACPs) to identify the genes that are specifically or prominently expressed in bovine early blastocysts and hatched blastocysts produced in vitro. Using these techniques, a total of nine expressed sequence tags (ESTs) of genes that were differentially expressed in hatched blastocysts, as compared to blastocyst embryos, were cloned and sequenced. The cloned genes or ESTs (C1-C9) all exhibited significant sequence similarity with known bovine genes (99-100%; FTL, RPS12, LAPTM4a, and RPL12) or ESTs (80-94%; AIBP, CULLIN-1, HDLP, COX5a, and RECS1) of other species. As revealed by real time RT-PCR, these genes were regulated upstream in the hatched blastocyst stage during early implantation. These results suggest that this new, PCR-based differential display RT-PCR technique is a very useful tool for the identification of stage-specific genes of preimplantation embryos.
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SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27
Andrea C. Carrano, Esther Eytan, Avram Hershko et al. · Nature Cell Biology · 1999 · 1.5K citations