Intervening sequence-specific in situ hybridization: detection of the pro-opiomelanocortin gene primary transcript in individual neurons

Robert T. Fremeau, Dominic J. Autelitano, Mariann Blum, Josiah Wilcox, James L. Roberts

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

We describe here the in situ hybridization procedure which we have used to detect the pro-opiomelanocortin (POMC) gene primary transcript in nuclei of individual neurons in the periarcuate region of the hypothalamus. An exon-intron RNA probe was used to detect POMC primary transcript and mature mRNA in nuclear extracts of nucleic acids using a sensitive S1 nuclease protection assay. The levels per cell of nuclear primary transcript were similar to those seen in the anterior pituitary, suggesting that intervening sequence in situ hybridization should be feasible. A nonrepetitive complementary RNA probe specific for the first intervening sequence of the rat POMC gene (POMC IVS-A) was used to detect the POMC primary transcript in hypothalamic tissue sections by in situ hybridization. The distribution of nuclear localized autoradiographic grains was similar to that previously reported for immunocytochemically defined POMC neurons, suggesting that the procedure is also effective in brain cells.

Original languageEnglish (US)
Pages (from-to)197-201
Number of pages5
JournalMolecular Brain Research
Volume6
Issue number2-3
DOIs
StatePublished - Nov 1989
Externally publishedYes

Keywords

  • Brain
  • In situ hybridization
  • Primary transcript
  • Pro-opiomelanocortin
  • Solution hybridization
  • mRNA

ASJC Scopus subject areas

  • Molecular Biology
  • Cellular and Molecular Neuroscience

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