TY - JOUR
T1 - Transcriptional activities of human elongation factor-1α and cytomegalovirus promoter in transgenic dogs generated by somatic cell nuclear transfer
AU - Eun, Kiyoung
AU - Hong, Nayoung
AU - Jeong, Yeon Woo
AU - Park, Min Gi
AU - Hwang, Seon Ung
AU - Jeong, Yeon I.K.
AU - Choi, Eun Ji
AU - Olsson, P. Olof
AU - Hwang, Woo Suk
AU - Hyun, Sang Hwan
AU - Kim, Hyunggee
N1 - Publisher Copyright:
© 2020 Eun et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2020/6/1
Y1 - 2020/6/1
N2 - Recent advances in somatic cell nuclear transfer (SCNT) in canines facilitate the production of canine transgenic models. Owing to the importance of stable and strong promoter activity in transgenic animals, we tested human elongation factor 1α (hEF1α) and cytomegalovirus (CMV) promoter sequences in SCNT transgenic dogs. After transfection, transgenic donor fibroblasts with the hEF1α-enhanced green fluorescence protein (EGFP) transgene were successfully isolated using fluorescence-activated cell sorting (FACS). We obtained four puppies, after SCNT, and identified three puppies as being transgenic using PCR analysis. Unexpectedly, EGFP regulated by hEF1α promoter was not observed at the organismal and cellular levels in these transgenic dogs. EGFP expression was rescued by the inhibition of DNA methyltransferases, implying that the hEF1α promoter is silenced by DNA methylation. Next, donor cells with CMV-EGFP transgene were successfully established and SCNT was performed. Three puppies of six born puppies were confirmed to be transgenic. Unlike hEF1α-regulated EGFP, CMV-regulated EGFP was strongly detectable at both the organismal and cellular levels in all transgenic dogs, even after 19 months. In conclusion, our study suggests that the CMV promoter is more suitable, than the hEF1α promoter, for stable transgene expression in SCNT-derived transgenic canine model.
AB - Recent advances in somatic cell nuclear transfer (SCNT) in canines facilitate the production of canine transgenic models. Owing to the importance of stable and strong promoter activity in transgenic animals, we tested human elongation factor 1α (hEF1α) and cytomegalovirus (CMV) promoter sequences in SCNT transgenic dogs. After transfection, transgenic donor fibroblasts with the hEF1α-enhanced green fluorescence protein (EGFP) transgene were successfully isolated using fluorescence-activated cell sorting (FACS). We obtained four puppies, after SCNT, and identified three puppies as being transgenic using PCR analysis. Unexpectedly, EGFP regulated by hEF1α promoter was not observed at the organismal and cellular levels in these transgenic dogs. EGFP expression was rescued by the inhibition of DNA methyltransferases, implying that the hEF1α promoter is silenced by DNA methylation. Next, donor cells with CMV-EGFP transgene were successfully established and SCNT was performed. Three puppies of six born puppies were confirmed to be transgenic. Unlike hEF1α-regulated EGFP, CMV-regulated EGFP was strongly detectable at both the organismal and cellular levels in all transgenic dogs, even after 19 months. In conclusion, our study suggests that the CMV promoter is more suitable, than the hEF1α promoter, for stable transgene expression in SCNT-derived transgenic canine model.
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U2 - 10.1371/journal.pone.0233784
DO - 10.1371/journal.pone.0233784
M3 - Article
C2 - 32492024
AN - SCOPUS:85085908576
VL - 15
JO - PLoS One
JF - PLoS One
SN - 1932-6203
IS - 6
M1 - e0233784
ER -