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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1979 Feb;76(2):625–629. doi: 10.1073/pnas.76.2.625

Parvovirus RNA transcripts containing sequences not present in mature mRNA: a method for isolation of putative mRNA precursor sequences.

F T Jay, L M de la Maza, B J Carter
PMCID: PMC383001  PMID: 218213

Abstract

We report here a method of RNA preparation that may enrich for precursor RNA sequences and the results of an investigation of adeno-associated virus (AAV) RNA transcription that used this method. Whole cells were lysed with detergent and high salt and separated into supernatant and pellet (crude chromatin) fractions. These fractions were then separately deproteinized by proteolytic digestion and phenol extractions. DNA was removed from the preparation by two cycles of pancreatic DNase digestion and phenol extraction. Hybridization analyses of the RNA obtained from AAV/adenovirus-infected KB (human) cells revealed some AAV-specific RNA sequences that were not present in the mature 20S mRNA. These additional sequences were contained in AAV RNA molecules present in the pellet fraction, whereas the 20S AAV mRNA accumulated in the supernatant. A species of AAV-specific RNA (about 22S), which was associated only with the pellet fraction and was labeled only after a short pulse, appeared to have a kinetic relationship with the more stable cytoplasmic 20S mRNA. These putative AAV mRNA "precursors" and precursor sequences were not observed previously when conventional methods were used to obtain RNA from either whole cells or isolated nuclei.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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