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Molecular Human Reproduction, Vol 3, 359-365, Copyright © 1997 by Oxford University Press

JOURNAL ARTICLE

Quantification of hexokinase mRNA in mouse blastocysts by competitive reverse transcriptase polymerase chain reaction

MD Johnson, DW Batey and B Behr
Department of Gynecology and Obstetrics, Stanford University School of Medicine, CA 94305-5317, USA.

Hexokinase (HX), the enzyme that catalyses the initial reaction in glycolysis, is an important enzyme in glucose metabolism during human and mouse embryonic development. In our previous investigations of the genetic activities of HX, we observed an increased incidence of HX gene expression in blastocysts in comparison with morulae, and variability in the incidence of HX gene expression between embryos at the same developmental stages. These observations prompted us to quantify HX mRNA in mouse blastocysts to define the biological significance of the variable gene transcription. We modified our qualitative reverse transcription-nested polymerase chain reaction (RT-nPCR) assay for HX mRNA in single or groups of embryos to quantify HX mRNA by competitive RT-nPCR. HX mRNA was quantified in cohorts of mouse blastocysts cultured in glucose/phosphate-containing human tubal fluid (HTF) media. These blastocysts expressed HX in minute amounts, averaging 1.95 x 10(- 18) g of mRNA. This is the first attempt at quantification of single gene mRNA in preimplantation embryos. Further investigations using similar techniques will enable comparative analyses between embryos to be performed to determine the correlation between specific levels of HX mRNA transcripts in individual embryos and embryonic viability and competence for further development and implantation.
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J. F.C. Chow, K.-F. Lee, S. T.H. Chan, and W. S.B. Yeung
Quantification of transforming growth factor {beta}1 (TGF{beta}1) mRNA expression in mouse preimplantation embryos and determination of TGF{beta} receptor (type I and type II) expression in mouse embryos and reproductive tract
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