Double in situ hybridization in combination with digital image analysis: a new approach to study interphase chromosome topography.
Double in situ hybridization with mercurated and biotinylated chromosome specific DNA probes in combination with digital image analysis provides a new approach to compare the distribution of homologous and nonhomologous chromosome targets within individual interphase nuclei. Here we have used two DNA probes representing tandemly repeated sequences specific for the constitutive heterochromatin of the human chromosomes 1 and 15, respectively, and studied the relative arrangements of these chromosome targets in interphase nuclei of human lymphocytes, amniotic fluid cells, and fibroblasts, cultivated in vitro. We have developed a 2D-image analysis approach which allows the rapid evaluation of large numbers of interphase nuclei. Models to test for a random versus nonrandom distribution of chromosome segments are discussed taking into account the three-dimensional origin of the evaluated 2D-distribution. In all three human diploid cell types the measurements of target-target and target-center distances in the 2D-nuclear image revealed that the labeled segments of the two chromosomes 15 were distributed both significantly closer to each other and closer to the center of the nuclear image than the labeled chromosome 1 segments. This result can be explained by the association of nucleolus organizer regions on the short arm of chromosome 15 with nucleoli located more centrally in these nuclei and does not provide evidence for a homologous association per se. In contrast, evaluation of the interphase positioning of the two chromosome 1 segments fits the random expectation in amniotic fluid and fibroblast cells, while in experiments using lymphocytes a slight excess of larger distances between these homologous targets was occasionally observed. 2D-distances between the labeled chromosome 1 and 15 segments showed a large variability in their relative positioning. In conclusion our data do not support the idea of a strict and permanent association of these homologous and nonhomologous targets in the cell types studied so far.
Emmerich
P
P
Institute of Anthropology and Human Genetics, University of Heidelberg, Federal Republic of Germany.
Loos
P
P
Jauch
A
A
Hopman
A H
AH
Wiegant
J
J
Higgins
M J
MJ
White
B N
BN
van der Ploeg
M
M
Cremer
C
C
Cremer
T
T
D002466:Cell Nucleolus / Q000648:ultrastructure; D002478:Cells, Cultured; D002900:Chromosomes, Human, 1-3 / Q000032:analysis / Q000648:ultrastructure*; D002901:Chromosomes, Human, 13-15 / Q000032:analysis / Q000648:ultrastructure*; D015342:DNA Probes; D003703:Demecolcine / Q000494:pharmacology; D005260:Female; D006570:Heterochromatin / Q000032:analysis*; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007399:Interphase; D008297:Male; D009693:Nucleic Acid Hybridization; D009997:Osmotic Pressure
D015342:DNA Probes; D006570:Heterochromatin; D003703:Demecolcine