Unknown,Transcriptomics,Genomics,Proteomics

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Global gene expression profiles in Oct4-knockdown and Ccna2-knockdown mouse embryos.


ABSTRACT: Gene regulation at the maternal-embryonic transition in the pre-implantation mouse embryo is not well understood. We knock down Ccna2 to establish proof-of-concept that antisense morpholino oligonucleotides can be used to target specific genes. We applied this strategy to study Oct4 and discovered that Oct4 is required prior to blastocyst development. Specifically, gene expression is altered as early as the 2-cell stage in Oct4-knockdown embryos. Distinct subsets of genes are differentially expressed between Oct4 and Ccna2-knockdown embryos, and indicated differential functions. Further, a large panel of genes were confirmed to be differentially-expressed in Oct4-knockdown embryos by quantitative, real time RT-PCR. Keywords: gene knockdown 3-5 week old wild type F1 (C57BL6xDBA/2) females (Charles River) were superovulated by intraperitonial injections of 5 IU of pregnant mare’s serum gonadotropin (Sigma) followed by 5 IU of human chorionic gonadotropin (Sigma) 48 hours later, and mated overnight with wild type males. Mice were sacrificed by cervical dislocation 17 hours after hCG injection, and 1-cell embryos were released from oviducts. Cumulus cells were removed by hyaluronidase (Sigma) treatment and pipetting. Pre-implantation embryos at the two pronuclei stage were recovered, pooled from 3-6 females in M2 media (Chemicon International), followed by immediate cytoplasmic microinjection of gene-specific antisense morpholino oligonucleotides and culture in Human Tubal Fluid with 10% serum supplement (In-Vitro Fertilization, Inc.) microdrops under mineral oil (Sigma) in mixed gas (90% nitrogen, 5% oxygen, 5% carbon dioxide; Praxair) at 37°C, and cultured at ten embryos per 20 μL drop. Uninjected control embryos derived from the same embryo pool, and were placed in identical conditions in parallel, except that they were not injected.

ORGANISM(S): Mus musculus

SUBMITTER: Bokyung Choi 

PROVIDER: E-GEOD-13981 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

A novel and critical role for Oct4 as a regulator of the maternal-embryonic transition.

Foygel Kira K   Choi Bokyung B   Jun Sunny S   Leong Denise E DE   Lee Alan A   Wong Connie C CC   Zuo Elizabeth E   Eckart Michael M   Reijo Pera Renee A RA   Wong Wing H WH   Yao Mylene W M MW  

PloS one 20081231 12


<h4>Background</h4>Compared to the emerging embryonic stem cell (ESC) gene network, little is known about the dynamic gene network that directs reprogramming in the early embryo. We hypothesized that Oct4, an ESC pluripotency regulator that is also highly expressed at the 1- to 2-cell stages in embryos, may be a critical regulator of the earliest gene network in the embryo.<h4>Methodology/principal findings</h4>Using antisense morpholino oligonucleotide (MO)-mediated gene knockdown, we show that  ...[more]

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