Molecular Human Reproduction, Vol. 5, No. 11, 1034-1039,
November 1999
© 1999 European Society of Human Reproduction and Embryology
Regulation of embryo development |
Analysis of gene expression in single oocytes and embryos by real-time rapid cycle fluorescence monitored RT–PCR
1 Gamete and Embryo Research Laboratory, Institute for Reproductive Medicine and Science of Saint Barnabas, West Orange, New Jersey, 07052, and 2 Department of Biology, Florida International University, Miami, Florida, 33199, USA
Abstract
Rapid cycle DNA amplification is a refinement of the polymerase chain reaction (PCR) method that permits increased product specificity while reducing amplification time by an order of magnitude. Combined with the use of micro volume capillaries, minute samples can be examined by this technique. Thus, this approach is ideally suited to the analysis of gene expression in individual cells. As the current understanding of early developmental processes is still rudimentary, further characterization of transcription in single oocytes and embryos may provide additional insight into the molecular mechanisms directing these events. In this study, we examined the suitability of fluorescence monitored reverse transcription (RT)–PCR for the study of gene expression during oogenesis and embryogenesis using transcripts of the housekeeping gene, ß-actin, as an experimental model. Product accumulation was monitored by either the double-stranded DNA dye SYBR Green I or sequence-dependent hybridization of reporter molecules called molecular beacons. Dyes bind generically and are economical to use. However, both specific and non-specific products are labelled. Hybridization probes permit very specific and sensitive target recognition but they can be costly to manufacture. Once molecular markers indicative of optimal development are identified, this technology could be used in a clinical in-vitro fertilization laboratory as a diagnostic tool.
human oocytes/molecular beacons/preimplantation embryos/rapid cycling/RT–PCR
Notes
3 To whom correspondence should be addressed
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  A. P. Gomez, M. J. Moreno, R. M. Baldrich, and A. Hernandez Endothelin-1 Molecular Ribonucleic Acid Expression in Pulmonary Hypertensive and Nonhypertensive Chickens Poult. Sci., July 1, 2008; 87(7): 1395 - 1401. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Sudheer and J. Adjaye Functional genomics of human pre-implantation development Brief Funct Genomic Proteomic, July 31, 2007; (2007) elm012v1. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. P. Gomez, M. J. Moreno, A. Iglesias, P. X. Coral, and A. Hernandez Endothelin 1, its Endothelin Type A Receptor, Connective Tissue Growth Factor, Platelet-Derived Growth Factor, and Adrenomedullin Expression in Lungs of Pulmonary Hypertensive and Nonhypertensive Chickens Poult. Sci., May 1, 2007; 86(5): 909 - 916. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 U. Eichenlaub-Ritter, U. Winterscheidt, E. Vogt, Y. Shen, H.-R. Tinneberg, and R. Sorensen 2-Methoxyestradiol Induces Spindle Aberrations, Chromosome Congression Failure, and Nondisjunction in Mouse Oocytes Biol Reprod, May 1, 2007; 76(5): 784 - 793. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. M. Steuerwald, M. D. Steuerwald, and J. B. Mailhes Post-ovulatory aging of mouse oocytes leads to decreased MAD2 transcripts and increased frequencies of premature centromere separation and anaphase Mol. Hum. Reprod., September 1, 2005; 11(9): 623 - 630. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Boiani, L. Gentile, V. V. Gambles, F. Cavaleri, C. A. Redi, and H. R. Scholer Variable Reprogramming of the Pluripotent Stem Cell Marker Oct4 in Mouse Clones: Distinct Developmental Potentials in Different Culture Environments Stem Cells, September 1, 2005; 23(8): 1089 - 1104. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. Wells, M.G. Bermudez, N. Steuerwald, A.R. Thornhill, D.L. Walker, H. Malter, J.D.A. Delhanty, and J. Cohen Expression of genes regulating chromosome segregation, the cell cycle and apoptosis during human preimplantation development Hum. Reprod., May 1, 2005; 20(5): 1339 - 1348. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Tranguch, N. Steuerwald, and Y. M. Huet-Hudson Nitric Oxide Synthase Production and Nitric Oxide Regulation of Preimplantation Embryo Development Biol Reprod, May 1, 2003; 68(5): 1538 - 1544. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. McGraw, C. Robert, L. Massicotte, and M.-A. Sirard Quantification of Histone Acetyltransferase and Histone Deacetylase Transcripts During Early Bovine Embryo Development Biol Reprod, February 1, 2003; 68(2): 383 - 389. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Robert, S. McGraw, L. Massicotte, M. Pravetoni, F. Gandolfi, and M.-A. Sirard Quantification of Housekeeping Transcript Levels During the Development of Bovine Preimplantation Embryos Biol Reprod, November 1, 2002; 67(5): 1465 - 1472. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Liss Improved quantitative real-time RT-PCR for expression profiling of individual cells Nucleic Acids Res., September 1, 2002; 30(17): e89 - e89. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J.-J. Hwang, P. D. Allen, G. C. Tseng, C.-W. Lam, L. Fananapazir, V. J. Dzau, and C.-C. Liew Microarray gene expression profiles in dilated and hypertrophic cardiomyopathic end-stage heart failure Physiol Genomics, July 12, 2002; 10(1): 31 - 44. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. W. Pfaffl, G. W. Horgan, and L. Dempfle Relative expression software tool (REST(C)) for group-wise comparison and statistical analysis of relative expression results in real-time PCR Nucleic Acids Res., May 1, 2002; 30(9): e36 - e36. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Jagannathan, E. L. Punt, Y. Gu, C. Arnoult, D. Sakkas, C. L. R. Barratt, and S. J. Publicover Identification and Localization of T-type Voltage-operated Calcium Channel Subunits in Human Male Germ Cells. EXPRESSION OF MULTIPLE ISOFORMS J. Biol. Chem., March 1, 2002; 277(10): 8449 - 8456. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. V. Hooper, M. H. Wong, A. Thelin, L. Hansson, P. G. Falk, and J. I. Gordon Molecular Analysis of Commensal Host-Microbial Relationships in the Intestine Science, February 2, 2001; 291(5505): 881 - 884. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Steuerwald, J. Cohen, R. J. Herrera, M. Sandalinas, and C. A. Brenner Association between spindle assembly checkpoint expression and maternal age in human oocytes Mol. Hum. Reprod., January 1, 2001; 7(1): 49 - 55. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. B. Shappell, R. A. Gupta, S. Manning, R. Whitehead, W. E. Boeglin, C. Schneider, T. Case, J. Price, G. S. Jack, T. M. Wheeler, et al. 15S-Hydroxyeicosatetraenoic Acid Activates Peroxisome Proliferator-activated Receptor {{gamma}} and Inhibits Proliferation in PC3 Prostate Carcinoma Cells Cancer Res., January 1, 2001; 61(2): 497 - 503. [Abstract] [Full Text]
|
|
|
|
|
|
M. Alikani, G. Calderon, G. Tomkin, J. Garrisi, M. Kokot, and J. Cohen Cleavage anomalies in early human embryos and survival after prolonged culture in-vitro Hum. Reprod., December 1, 2000; 15(12): 2634 - 2643. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. H. Wong, J. R. Saam, T. S. Stappenbeck, C. H. Rexer, and J. I. Gordon Genetic mosaic analysis based on Cre recombinase and navigated laser capture microdissection PNAS, October 23, 2000; (2000) 230237997. [Abstract] [Full Text]
|
|
|
|
|
|
N. Steuerwald, J. Cohen, R. J. Herrera, and C. A. Brenner Quantification of mRNA in single oocytes and embryos by real-time rapid cycle fluorescence monitored RT-PCR Mol. Hum. Reprod., May 1, 2000; 6(5): 448 - 453. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. H. Wong, J. R. Saam, T. S. Stappenbeck, C. H. Rexer, and J. I. Gordon Genetic mosaic analysis based on Cre recombinase and navigated laser capture microdissection PNAS, November 7, 2000; 97(23): 12601 - 12606. [Abstract] [Full Text] [PDF]
|
|