Gene expression profiling of mouse oocytes and preimplantation embryos.
Gene expression profiling using microarray technology is a robust, efficient, and cost-effective approach to assay a cell or tissue's transcriptome at a particular time or under a specific condition. Application of this technology to oocytes and preimplantation embryos has been limited largely because this biological material is difficult to acquire in sufficient quantities. We describe here a protocol to isolate and amplify mRNA from oocytes and preimplantation embryos that is suitable for microarray experiments. This protocol is based on a linear two-step amplification protocol using T7 RNA polymerase-based in vitro transcription and has been used to isolate more than 80 microg of cRNA from only 20 oocytes or preimplantation embryos. Gene expression profiling has provided insight into the molecular mechanisms of meiotic maturation, fertilization, and preimplantation embryo development. It has also been used to characterize female gametes and embryos from animals harboring gene-specific knockouts or knockdowns. Finally, this technology has been useful in evaluating how various Assisted Reproductive Technologies impact global patterns of gene expression in resulting embryos.
Duncan
Francesca E
FE
Department of Obstetrics and Gynecology, Northwestern University, Chicago, Illinois, USA.
Schultz
Richard M
RM
D000818:Animals; D001755:Blastocyst / Q000378:metabolism*; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods*; D051379:Mice; D009865:Oocytes / Q000378:metabolism*; D011247:Pregnancy