The addition of serum to embryo culture media may alter gene expression and trigger development of Large Offspring Syndrome. The objectives of this study were to determine gene expression levels in embryos cultured in the absence or presence of 5% calf serum and compare these expression patterns to in vivo derived embryos (IVD), and to determine the effects of serum on the length of day-14 embryos. Abattoir derived oocytes were fertilized and cultured in mSOFaa. At 72 hours post-insemination (hpi), embryos were randomly allocated into two treatments: mSOFaa without and with 5% calf serum. Embryos were then cultured to 168 hpi and blastocyst rates were assessed. In experiment 1, blastocysts from each treatment were pooled and stored at -80°C. In experiment 2, blastocysts (n=5-10) from each treatment were transferred into synchronized recipients, and were recovered 7 days post-transfer. Embryos were photographed, measured, and immediately stored at -80°C. Isolation of mRNA, reverse transcription and quantitative PCR were performed to determine transcript abundance for COX6A, IFNT1a, PLAC8, IGF2R and GAPDH for each sample. In both experiments, blastocyst development rates were higher in embryos cultured with serum compared to the no-serum treatment (14.9 and 7.4% respectively, P<0.001). In experiment 1, no differences were found in the expression of COX6A, IFNT1a, IGF2R and PLAC8; however upregulated expression of IGF2R, COX6A and IFNT1a were observed in some samples in both IVP treatments. In experiment 2, lengths of elongated embryos from the serum and no-serum culture treatments differed from the IVD treatment. Mean expression levels for COX6A, IFNT1a, PLAC8 and IGF2R did not differ across treatment groups. However, in the serum treatment 3 of 11 embryos over-expressed IFNT1a, 4 of 11 over-expressed IGF2R and 2 of 11 over-expressed PLAC8, over-expression being defined as two standard deviations above the mean of the IVD treatment for each respective gene. While mean expression levels were not affected by culture with serum under these conditions, very high expression of IFNT1a, IGF2R and PLAC8 in experiment 2 and IGF2R and IFNT1a in experiment 1 was observed in some embryos cultured with serum, but not in embryos cultured without serum or in in vivo derived embryos.
| Identifer | oai:union.ndltd.org:LSU/oai:etd.lsu.edu:etd-11122010-115305 |
| Date | 17 November 2010 |
| Creators | Angulo Campos, Jaime Manuel |
| Contributors | Godke, Robert, Bondioli, Kenneth, Lyle, Sara, Gentry, Glen |
| Publisher | LSU |
| Source Sets | Louisiana State University |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.lsu.edu/docs/available/etd-11122010-115305/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached herein a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to LSU or its agents the non-exclusive license to archive and make accessible, under the conditions specified below and in appropriate University policies, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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