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Identification of differentially regulated genes in bovine blastocysts using an annealing control primer system
Author(s) -
Hwang KyuChan,
Cui XiangShun,
Park SePill,
Shin MiRa,
Park SaeYoung,
Kim EunYoung,
Kim NamHyung
Publication year - 2004
Publication title -
molecular reproduction and development
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 105
eISSN - 1098-2795
pISSN - 1040-452X
DOI - 10.1002/mrd.20156
Subject(s) - biology , primer (cosmetics) , gene , genetics , computational biology , identification (biology) , chemistry , organic chemistry , botany
The identification of embryo‐specific genes would provide insights into early embryonic development. However, the current methods employed to identify the genes that are expressed at a specific developmental stage are labor intensive and suffer from high rates of false positives. Here we employed a new and accurate reverse transcription‐polymerase chain reaction (RT‐PCR) technology that involves annealing control primers (ACPs) to identify the genes that are specifically or prominently expressed in bovine early blastocysts and hatched blastocysts produced in vitro. Using these techniques, a total of nine expressed sequence tags (ESTs) of genes that were differentially expressed in hatched blastocysts, as compared to blastocyst embryos, were cloned and sequenced. The cloned genes or ESTs (C1–C9) all exhibited significant sequence similarity with known bovine genes (99–100%; FTL, RPS12, LAPTM4a, and RPL12) or ESTs (80–94%; AIBP, CULLIN‐1, HDLP, COX5a, and RECS1) of other species. As revealed by real time RT‐PCR, these genes were regulated upstream in the hatched blastocyst stage during early implantation. These results suggest that this new, PCR‐based differential display RT‐PCR technique is a very useful tool for the identification of stage‐specific genes of preimplantation embryos. Mol. Reprod. Dev. 69: 43–51, 2004. © 2004 Wiley‐Liss, Inc.