Polytene chromosome squash methods for studying transcription and epigenetic chromatin modification in Drosophila using antibodies.
The giant polytene chromosomes from Drosophila third instar larval salivary glands provide an important model system for studying the architectural changes in chromatin morphology associated with the process of transcription initiation and elongation. Especially, analysis of the heat shock response has proved useful in correlating chromatin structure remodeling with transcriptional activity. An important tool for such studies is the labeling of polytene chromosome squash preparations with antibodies to the enzymes, transcription factors, or histone modifications of interest. However, in any immunohistochemical experiment there will be advantages and disadvantages to different methods of fixation and sample preparation, the relative merits of which must be balanced. Here we provide detailed protocols for polytene chromosome squash preparation and discuss their relative pros and cons in terms of suitability for reliable antibody labeling and preservation of high resolution chromatin structure.
Johansen
Kristen M
KM
Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, 3154 Molecular Biology Building, Ames, IA 50011, USA. kristen@iastate.edu
Cai
Weili
W
Deng
Huai
H
Bao
Xiaomin
X
Zhang
Weiguo
W
Girton
Jack
J
Johansen
Jørgen
J
D000818:Animals; D002843:Chromatin / Q000235:genetics / Q000502:physiology*; D002875:Chromosomes / Q000737:chemistry; D004330:Drosophila; D044127:Epigenesis, Genetic; D007124:Immunoenzyme Techniques; D014158:Transcription, Genetic
D002843:Chromatin