Expression of the recombinase-activating gene (RAG-1) in murine early embryogenesis.
The recombinase activation genes, RAG-1 and RAG-2, are expressed together in immature T or B lymphocytes and possess activity to induce V(D)J rearrangement in T cell receptor (TCR) and Ig genes. In vertebrates, only Ig and TCR molecules are reported to have recombination in their development using multiple V, D, J component gene segments. Thus, expression of RAG genes are localized only in lymphoid organs and sites of extrathymic T cell differentiation. In this study, we have used RAG-1 and RAG-2 genes as markers of possible genetic recombination in developing murine preimplantation embryos, using the highly sensitive reverse transcriptase polymerase chain reaction (RT-PCR) technique and in situ hybridization. From 40 preimplantation embryos of various developmental stages we extracted RNA, reverse-transcribed it into cDNA and used it in RT-PCR studies. A PCR of 35 cycles disclosed expression of RAG-1 but not RAG-2 in morulae and blastocysts. Southern blot hybridization using a specific synthetic oligonucleotide probe for RAG-1 and RT-PCR with another primer pair identified RAG-1 expression in developing embryos. In situ hybridization using a cooled CCD camera also revealed localization of RAG-1 mRNA in blastocysts. We propose possible genetic recombination during late preimplantation murine embryogenesis which may contribute to the loss of totipotency and differentiation of inner cell mass and trophoectoderm.
Hayakawa
S
S
Department of Obstetrics and Gynaecology, Nihon University, Tokyo, Japan.
Tochigi
M
M
Chishima
F
F
Shiraishi
H
H
Takahashi
N
N
Watanabe
K
K
Fujii
K T
KT
Satoh
K
K
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D015870:Gene Expression; D016654:Genes, RAG-1; D017403:In Situ Hybridization; D051379:Mice; D008813:Mice, Inbred ICR / Q000196:embryology / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
D018076:DNA, Complementary