Rapid, high-resolution in situ hybridization histochemistry with radioiodinated synthetic oligonucleotides

Journal Article · Wed Jan 01 00:00:00 EST 1986 · J. Neurosci. Res.; (United States)

DOI:https://doi.org/10.1002/jnr.490160112· OSTI ID:5009044

Lewis, M E; Arentzen, R; Baldino, F Jr

In situ hybridization histochemistry is a valuable technique for localizing specific messenger RNA (mRNA) and detecting changes in gene expression. Generally, the mRNA of interest has been detected by probes obtained from cloned DNA and labelled to high specific activity by nick translation. Such probes have a number of disadvantages which can be circumvented by the use of short synthetic oligonucleotides designed to be complementary to a known mRNA sequence. We report here that synthetic oligonucleotides complementary to part of the mRNA coding for rat arginine-vasopressin (AVP) can be labelled to high specific activity with (/sup 125/I), using either the primer extension method with the Klenow fragment of DNA polymerase I or the 3'-tailing method with terminal deoxynucleotidyl transferase. Both AVP probes hybridized well to the magnocellular neurons of the hypothalamic paraventricular and supraoptic nuclei. A strong autoradiographic signal was present by 2 days, with grains largely confined to the perikaryon. These results compare favorably to those obtained with (/sup 32/P)- or (/sup 3/H)-labelled probes. Given the ease of the 3'-tailing method, (/sup 125/I)-labelled oligonucleotides appear to be especially useful probes for in situ hybridization histochemistry.

• Research Organization: E.I. du Pont de Nemours and Co., Wilmington, DE
• OSTI ID: 5009044
• Journal Information: J. Neurosci. Res.; (United States), Vol. 1
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
MESSENGER-RNA
AUTORADIOGRAPHY
NUCLEOTIDES
DNA
HISTOLOGICAL TECHNIQUES
HYPOTHALAMUS
IODINE 125
PHOSPHORUS 32
RATS
TRITIUM COMPOUNDS
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BODY
BRAIN
CENTRAL NERVOUS SYSTEM
DAYS LIVING RADIOISOTOPES
ELECTRON CAPTURE RADIOISOTOPES
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPES
LABELLED COMPOUNDS
LIGHT NUCLEI
MAMMALS
NERVOUS SYSTEM
NUCLEI
NUCLEIC ACIDS
ODD-EVEN NUCLEI
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANS
PHOSPHORUS ISOTOPES
RADIOISOTOPES
RNA
RODENTS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques