Telomerase activity and telomere detection during early bovine development.
The ends of mammalian chromosomes are composed of repeated DNA sequences of (TTAGGG)(n) known as telomeres. Telomerase is a ribonucleoprotein that synthesizes telomeric DNA to replenish the 50-200 bp lost during cell replication. Cellular aging and senescence are associated with a lack of telomerase activity and a critical shortening of the telomere. The objectives of this study were to confirm the presence of TTAGGG repeats on the chromosomes of bovine embryos using in situ hybridization and assess the relative amounts of telomerase activity using a telomeric repeat amplification protocol (TRAP) during oocyte maturation and early embryo development. Applying a telomere DNA probe to the chromosomes of blastocysts and adult fibroblasts, telomeres were identified on the terminal ends of the p and q arms of chromosomes in all cells examined. Immature oocytes, matured oocytes, zygotes, 2- to 5-cell embryos, 6- to 8-cell embryos, morulae, and blastocysts were lysed in NP-40 lysis buffer and telomerase activity was assayed using the TRAP assay. Telomerase activity was detected in all developmental stages examined. Relative telomerase activity (based on telomerase internal standards and positive controls) appeared to decrease during oocyte maturation and subsequent development to the 8-cell stage but significantly increased (P < 0.05) by approximately 40-fold at the morula and blastocyst stages. It was concluded that the telomeres of bovine chromosomes contain TTAGGG repeats and that telomerase activity is up-regulated in morulae and blastocysts.
Betts
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DH
Department of Biomedical Sciences, Ontario Veterinary College, University of Guelph, Guelph, Ontario, Canada.
King
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WA
D000818:Animals; D002417:Cattle / Q000196:embryology / Q000378:metabolism*; D015342:DNA Probes; D005307:Fertilization in Vitro; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D009865:Oocytes / Q000254:growth & development / Q000378:metabolism*; D020080:Tandem Repeat Sequences / Q000235:genetics; D019098:Telomerase / Q000378:metabolism*; D016615:Telomere / Q000235:genetics*
D015342:DNA Probes; D019098:Telomerase