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Extra info for Chromatin Structure and Function: Molecular and Cellular Biophysical Methods

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Wilson, R. & Bonner, J. (1974) Proc. Nat. Acad. Sci. ]1:2193-2197. , Berg, B. & Bonner, J. (1976) Biochemistry 15 : 2742-2482 . Gare1, A. & Axel, R. (1976) Proc. Nat. Acad. Sci. 11:3966-3970. Weintraub, H. & Groudine, H. (1976) Science 193:848-856. , Radda, G. & Walker, I. o. (1974) Biochemistry 13:2937-2945. , Miller, o. & Bakken, A. (1973) Cold Spring Harbor Symp. on Quant. Bio1. 38:915-919. ~ Mirsky, A. E. (1964) Proc. Nat. Acad. Sci. 11:3937-3941. Marushige, K. (1976) Proc. Nat. Acad. Sci.

A demonstration of differential gene expression requires evidence for a selection of DNA sequences transcribed into RNA. , 1968) and mouse liver uterus before and after oestrogen stimulation (Church and McCarthy, 1970). The finding that eukaryotic chromatin can support the transcription of RNA in vitro in the presence of added RNA polymerase, usually bacterial, suggested a cell-free approach to the study of this phenomenon. Us ing the same hybridization techniques to compare the homologies of in vitro transcribed RNA and total genomic DNA it was concluded that~ability of chromatin to support RNA synthesis is restricted compared with that of DNA.

In a control incubation RNA polymerase was omitted and an appropriate amount of t-RNA added. Re-isolation of the t-RNA was carried out in parallel with the transcripts; significant hybridisation was seen in this case. The globin RNA sequences deteoted in the oontrol have come from the endogenous RNA that co-isolates with the chromatin and oontributes an estimated 50% of the observed hybridisation in aotively transcribing inoubations. The most serious criticism for the present argument is that any in vivo synthesized endogenous RNA isolated with the chromatin template will be indistinguishable from in vitro RNA synthesized by the exogenously added po~erase unl8ss-careful controls are included.

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