[Use of the modified method of chromatin immunoprecipitation for the isolation of actively transcribed loci].
A modified version of the chromosomal immunoprecipitation (ChIP) assay was implemented for discrete isolation and characterization of actively transcribed genes. Specifically, it was demonstrated with the gene II/9-1 of Sciara coprophila as a model locus that significant enhancement in the isolation of actively transcribed versus repressed and inactive genes can be achieved through the ChIP methodology. A combination of solid-phase magnetic bead technology with chromosomal immunoprecipitation using antibodies that recognize the large subunit (c) of RNA polymerase II resulted in efficient isolation of the promoter region of gene II/9-1 exclusively during the amplification stage of larval development, when the gene is actively transcribed. It is postulated that the novel technology described herein can be applied to a wide variety of systems for efficient isolation and in vivo assessment of actively transcribed genes regulated by virtually any given transcription factor.
Luniak
V
V
Petersburg Nuclear Physics Institute, Molecular and Radiation Physics Division, St. Petersburg, 188350 Russia.
Burgess
R
R
Noskin
L
L
Gil'iano
N
N
D000818:Animals; D002843:Chromatin / Q000737:chemistry*; D017931:DNA Primers; D004175:Diptera / Q000235:genetics / Q000254:growth & development; D005557:Formaldehyde / Q000737:chemistry; D015870:Gene Expression; D017344:Genes, Insect; D007814:Larva; D016133:Polymerase Chain Reaction / Q000379:methods; D011233:Precipitin Tests / Q000379:methods*; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000276:immunology; D014158:Transcription, Genetic
D002843:Chromatin; D017931:DNA Primers; D005557:Formaldehyde; D012319:RNA Polymerase II