An Artifactual In Situ Hybridization Signal Associated with Apoptosis in Rat Embryo

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Abstract
We report an artifactual in situ hybridization (ISH) labeling pattern in embryonic rat tissues. It is caused by a short multiple cloning site-derived sequence incorporated into the RNA probes by in vitro transcription of templates cloned into pBluescript or its descendants. The artifact was seen in tissues in which programmed cell death (apoptosis) takes place during embryogenesis, i.e., in the mesonephric area, developing nervous system, interdigital mesenchyme of the hand plate, and permanent kidney. Labeling of the radioactive ISH with TUNEL verified the co-localization of the artifactual hybridization signal with cells at early stages of apoptosis. Even though the identity of the hybridization target in apoptotic cells remains unknown, it might be highly species-specific, because this artifact was never observed in mouse tissues.