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The objective of this study was to examine the time during
the postfertilization period that gene expression patterns
in in vitro cultured bovine embryos diverge from those of
their in vivo cultured counterparts. Presumptive bovine
zygotes were produced by in vitro maturation and
fertilization of immature oocytes collected from the
ovaries of slaughtered animals. Approximately 20 h post
insemination (hpi), zygotes were denuded and randomly
divided in two groups for culture either in vitro, in
synthetic oviduct fluid medium (SOF), or in vivo, in the
ewe oviduct. Embryos were recovered from both systems at
approximately 30 hpi (2-cell), 2 (4-cell), 3 (8-cell), 4
(16-cell), 5 (early morula), 6 (compact morula) or 7
(blastocyst) days pi. On recovery, they were examined for
stage of development and snap frozen in liquid nitrogen
for the analysis of transcript abundance using real-time
PCR. The transcripts studied were glucose transporter 5,
sarcosine oxidase, mitochondrial Mn-superoxide dismutase,
connexin 43, interferon tau, insulin-like growth factor
II, apoptosis regulator box-
and insulin-like
growth factor-I receptor, most of which are known from our
previous work to differ in terms of transcript abundance
in blastocysts derived from culture in vitro or in vivo.
The results demonstrate that the relative abundance of the
transcripts studies varies throughout the preimplantation
period and is strongly influenced by the culture
environment. In addition, the data demonstrate that
changes in transcript abundance in blastocyst stage
embryos are in many cases a consequence of perturbed
transcription earlier in development. Depending on the
transcript these differences may be evident by as little
as 10 h of initiation of culture. Such information has
implications not only for basic biology but also for human
assisted reproduction where there is a move towards
culturing embryos to the blastocyst stage, necessitating
prolonged culture in vitro under potentially deleterious
conditions.
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