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Analysis of G protein α subunit mRNA abundance in preimplantation mouse embryos using a rapid, quantitative RT‐PCR approach
Author(s) -
Rambhatla Lakshmi,
Patel Bela,
Dhanasekaran N.,
Latham Keith E.
Publication year - 1995
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.1080410306
Subject(s) - biology , blastocyst , messenger rna , embryo , microbiology and biotechnology , protein subunit , complementary dna , population , alpha (finance) , reverse transcription polymerase chain reaction , real time polymerase chain reaction , embryogenesis , gene , genetics , medicine , construct validity , demography , nursing , sociology , patient satisfaction
We have developed a novel reverse transcription‐polymerase chain reaction (RT‐PCR)‐based approach for systematically quantifying in a single experiment the abundances of many different mRNAs in preimplantation mouse embryos. With this approach, the entire mRNA population from a small number of embryos is amplified while preserving the relative abundance of each mRNA in the cDNA population. The cDNA is analyzed by quantitative hybridization to radiolabeled probes. The approach is very sensitive and provides reliable, quantitative data regarding changes in mRNA abundance. A major advantage of this method is that estimates of mRNA copy number can be obtained and compared between different mRNAs. With this approach, we analyzed the patterns of expression of nine G protein α subunit mRNAs (Gα s , Gα i , Gα q′ , Gα o′ , and Gα 11–15 ) in oocytes, eggs, and preimplantation embryos from fertilization to the blastocyst stage. Six α subunit mRNAs were expressed at significant levels, all of which underwent significant temporal alterations in expression. The mRNAs encoding some α subunit types were expressed predominantly in the egg and 1‐cell embryo, underwent sharp reductions during the 2‐cell stage, and were re‐expressed between the 8‐cell and blastocyst stages. One α subunit mRNA increased in abundance at the early blastocyst stage. The possible significance of these alterations in G protein mRNA abundance to embryonic development is discussed. © 1995 Wiley‐Liss, Inc.