Analysis of development-related gene expression in cloned bovine blastocysts with different developmental potential.
The high incidence of abnormalities in cloned calves is a most serious problem for bovine somatic cell nuclear transfer (NT) technology. Because there is little information on the differences in mRNA expression in cloned blastocysts with donor cells of different sex and origin, we compared development-related gene expression in two types of cloned bovine blastocysts with different potentials to develop into normal calves, a female adult cumulus cell line (high potential to develop into live calves) and a male fibroblast cell line (low potential to develop into live calves) to examine the correlation between the normality of cloned calves and blastocyst mRNA expression patterns. We analyzed 12 genes involved in apoptosis, growth factor signaling, metabolism, and DNA methylation in blastocysts originating from two types of donor cells and in vitro-fertilized blastocysts using quantitative real-time polymerase chain reaction. Expression of the pro-apoptotic Bax gene and anti-apoptotic Bcl-2 and Glut-1 genes in fibroblast-derived blastocysts was significantly higher than in cumulus cell-derived and in vitro-fertilized blastocysts. The high Bcl-2 and Glut-1 gene expression suggests that some embryonic cells with damaged DNA in fibroblast-derived blastocysts are not removed, and their descendants later manifest abnormal placenta or fetus formation. Transfer of pre-selected cloned blastocysts into recipients is required, however, to determine whether the expression pattern of these apoptosis-related genes reflects differences in the potential to develop into normal calves.
Laboratory of Animal Reproduction, College of Agriculture, Kinki University, Nakamachi, Nara, Japan.
Xiangping
X
Li
Dasari
D
Amarnath
Yoko
Y
Kato
Yukio
Y
Tsunoda
D000818:Animals; D017209:Apoptosis; D001755:Blastocyst / Q000737:chemistry / Q000502:physiology*; D002417:Cattle; D002460:Cell Line; D002478:Cells, Cultured; D019976:Cloning, Organism / Q000379:methods*; D004247:DNA / Q000378:metabolism; D047108:Embryonic Development; D005260:Female; D005347:Fibroblasts / Q000737:chemistry / Q000166:cytology / Q000502:physiology; D018507:Gene Expression Regulation, Developmental; D051272:Glucose Transporter Type 1 / Q000032:analysis / Q000235:genetics; D008297:Male; D053652:Nuclear Transfer Techniques; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000032:analysis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D051028:bcl-2-Associated X Protein / Q000032:analysis / Q000235:genetics
D051272:Glucose Transporter Type 1; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051028:bcl-2-Associated X Protein; D004247:DNA