Analysis of the size distribution of first-strand cDNA molecules.
We have developed a method to analyze the size distribution of the first-strand cDNA molecules corresponding to given mRNA species. First-strand molecules synthesized from cytoplasmic polyadenylated RNAs are separated by electrophoresis on an alkaline agarose gel, and a Southern blot hybridization is performed. As an example, we analyzed the first-strand molecules corresponding to the human c-myc mRNAs. This method can be used to determine whether full-length, first-strand molecules corresponding to an mRNA species to be cloned are synthesized efficiently. Interestingly, this method allows one to analyze full-length, first-strand cDNA molecules with a much higher resolution than Northern blot analysis of mRNA molecules. This method can therefore be used to discriminate between the multiple mRNA species transcribed from a given gene or the homologous mRNA species transcribed from a given gene family.
Bodescot
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M
Institut Gustave-Roussy Villejuif, France.
Brison
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D015139:Blotting, Southern; D003001:Cloning, Molecular; D015342:DNA Probes; D018076:DNA, Complementary / Q000032:analysis* / Q000096:biosynthesis; D004587:Electrophoresis, Agar Gel; D016259:Genes, myc; D006801:Humans; D009693:Nucleic Acid Hybridization; D011061:Poly A / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics*; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D016914:Ribonuclease H / Q000378:metabolism; D014158:Transcription, Genetic
D015342:DNA Probes; D018076:DNA, Complementary; D012333:RNA, Messenger; D011061:Poly A; D012194:RNA-Directed DNA Polymerase; D016914:Ribonuclease H