TY - JOUR
T1 - Cytoplasmic DNA can be detected by RNA fluorescence in situ hybridization
AU - Greenberg, Eliraz
AU - Hochberg-Laufer, Hodaya
AU - Blanga, Shalev
AU - Kinor, Noa
AU - Shav-Tal, Yaron
N1 - Publisher Copyright: © The Author(s) 2019
PY - 2019/10/10
Y1 - 2019/10/10
N2 - Fluorescence in situ hybridization (FISH) can be used for the intracellular detection of DNA or RNA molecules. The detection of DNA sequences by DNA FISH requires the denaturation of the DNA double helix to allow the hybridization of the fluorescent probe with DNA in a single stranded form. These hybridization conditions require high temperature and low pH that can damage RNA, and therefore RNA is not typically detectable by DNA FISH. In contrast, RNA FISH does not require a denaturation step since RNA is single stranded, and therefore DNA molecules are not detectable by RNA FISH. Hence, DNA FISH and RNA FISH are mutually exclusive. In this study, we show that plasmid DNA transiently transfected into cells is readily detectable in the cytoplasm by RNA FISH without need for denaturation, shortly after transfection and for several hours. The plasmids, however, are usually not detectable in the nucleus except when the plasmids are efficiently directed into the nucleus, which may imply a more open packaging state for DNA after transfection. This detection of plasmid DNA in the cytoplasm has implications for RNA FISH experiments and opens a window to study conditions when DNA is present in the cytoplasm.
AB - Fluorescence in situ hybridization (FISH) can be used for the intracellular detection of DNA or RNA molecules. The detection of DNA sequences by DNA FISH requires the denaturation of the DNA double helix to allow the hybridization of the fluorescent probe with DNA in a single stranded form. These hybridization conditions require high temperature and low pH that can damage RNA, and therefore RNA is not typically detectable by DNA FISH. In contrast, RNA FISH does not require a denaturation step since RNA is single stranded, and therefore DNA molecules are not detectable by RNA FISH. Hence, DNA FISH and RNA FISH are mutually exclusive. In this study, we show that plasmid DNA transiently transfected into cells is readily detectable in the cytoplasm by RNA FISH without need for denaturation, shortly after transfection and for several hours. The plasmids, however, are usually not detectable in the nucleus except when the plasmids are efficiently directed into the nucleus, which may imply a more open packaging state for DNA after transfection. This detection of plasmid DNA in the cytoplasm has implications for RNA FISH experiments and opens a window to study conditions when DNA is present in the cytoplasm.
UR - http://www.scopus.com/inward/record.url?scp=85072718469&partnerID=8YFLogxK
U2 - https://doi.org/10.1093/NAR/GKZ645
DO - https://doi.org/10.1093/NAR/GKZ645
M3 - مقالة
C2 - 31340014
SN - 0305-1048
VL - 47
SP - E109
JO - Nucleic acids research
JF - Nucleic acids research
IS - 18
ER -