原创力文档selection of stable reference genes for quantitative rt-pcr comparisons of mouse embryonic and extra-embryonic stem cells选择稳定的参考基因定量rt - pcr比较小鼠胚胎干细胞和胚外.pdfVIP
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selection of stable reference genes for quantitative rt-pcr comparisons of mouse embryonic and extra-embryonic stem cells选择稳定的参考基因定量rt - pcr比较小鼠胚胎干细胞和胚外
Selection of Stable Reference Genes for Quantitative RT-
PCR Comparisons of Mouse Embryonic and Extra-
Embryonic Stem Cells
Kylee J. Veazey, Michael C. Golding*
College of Veterinary Medicine and Biomedical Sciences, Texas AM University, College Station, Texas, United States of America
Abstract
Isolation and culture of both embryonic and tissue specific stem cells provide an enormous opportunity to study the
molecular processes driving development. To gain insight into the initial events underpinning mammalian embryogenesis,
pluripotent stem cells from each of the three distinct lineages present within the preimplantation blastocyst have been
derived. Embryonic (ES), trophectoderm (TS) and extraembryonic endoderm (XEN) stem cells possess the developmental
potential of their founding lineages and seemingly utilize distinct epigenetic modalities to program gene expression.
However, the basis for these differing cellular identities and epigenetic properties remain poorly defined. Quantitative
reverse transcription-polymerase chain reaction (qPCR) is a powerful and efficient means of rapidly comparing patterns of
gene expression between different developmental stages and experimental conditions. However, careful, empirical
selection of appropriate reference genes is essential to accurately measuring transcriptional differences. Here we report the
quantitation and evaluation of fourteen commonly used references genes between ES, TS and XEN stem cells. These
included: Actb, B2m, Hsp70, Gapdh, Gusb, H2afz, Hk2, Hprt, Pgk1, Ppia, Rn7sk, Sdha, Tbp and Ywhaz. Utilizing three
independent statistical analysis, we identify Pgk1, Sdha and Tbp as the most stable reference genes between each of these
stem cell types. Furthermore, we identify Sdha, Tbp and Ywha
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