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Nucleic Acids Research, 1990, Vol. 18, No. 10 2987-2992
© 1990

MOLECULAR BIOLOGY

Regulation of kappa immunoglobulin gene transcription in vitro

R. Alexander Currie

Department of Medicine, Biochemistry, Genetics and Biophysics, University of Colorado Health Sciences Center Denver, CO 80262, USA

Received December 4, 1989. Accepted April 9, 1990.

An in vitro transcription system has been established with a whole-cell extract from the human Burkitt's lymphoma, Daudi, cell line. The B cell extract has been compared with a HeLa cell extract in an effort to study lymphocyte-specific regulatory factors of kappa light chain gene transcription. Both extracts were capable of transcribing Vk genes and other RNA polymerase II dependent genes. Alpha-amanitin at [0.1 /tg/ml] completely inhibited the accumulation of transcripts in both systems. At low DNA template concentrations the kappa intronic enhancer stimulated Vk promoter transcription 3–7 fold in B cell extracts. The enhancerdependent transcription was abolished by excising the enhancer from the test plasmid with Eco R1. Both Vk promoter and enhancer-dependent transcription in HeLa extracts was undetectable at low [DNA]. These results demonstrate that kappa enhancer stimulation of Vk transcription in vitro is observed in B cell extracts only under conditions of low DNA template concentration.


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