A comparative study of gene expression in murine embryos developed in vivo, cultured in vitro, and cocultured with human oviductal cells using messenger ribonucleic acid differential display.
The objectives of this study were to compare the mRNA expression patterns in early mouse embryos in different culture conditions by differential display reverse transcription-polymerase chain reaction (DDRT-PCR). Embryos developed in vivo, cultured in vitro, and cocultured with human oviductal epithelial cells were studied at the 2-cell, 4-cell, 8-cell/morula, and blastocyst stages. Messenger RNA profiles were displayed by DDRT-PCR using downstream T11VV (V = A, C, or G) and upstream decamer primers. Total cDNA banding patterns were highly conserved in the three groups studied. Some fragments are unique in different culture conditions. Thirteen out of the 40 selected differentially expressed clones were characterized. The DNA sequence analyses of these clones displayed high sequence homology with cDNA sequences in the mouse expressed sequence tag database. Using semiquantitative RT-PCR, we confirmed differential expression of these DD amplicons in the three groups of embryos. The temporal expression of some of the selected DD amplicons during preimplantation development were studied in the three groups of embryos. In conclusion, DDRT-PCR is an effective tool for contrasting gene expression patterns and characterizing mRNA transcripts in mouse embryo.
Lee
K F
KF
Department of Obstetrics and Gynaecology, Queen Mary Hospital, and Department of Zoology, The University of Hong Kong, Hong Kong, China.
Chow
J F
JF
Xu
J S
JS
Chan
S T
ST
Ip
S M
SM
Yeung
W S
WS
D000818:Animals; D003001:Cloning, Molecular; D018920:Coculture Techniques; D004247:DNA / Q000737:chemistry; D017931:DNA Primers / Q000737:chemistry; D005314:Embryonic and Fetal Development / Q000235:genetics*; D004847:Epithelial Cells / Q000166:cytology; D005187:Fallopian Tubes / Q000166:cytology*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D011247:Pregnancy; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
D017931:DNA Primers; D012333:RNA, Messenger; D004247:DNA