Transcription in vitro of immunoglobulin kappa light chain genes in isolated mouse myeloma nuclei and chromatin

Abstract

Messenger RNA sequences for immunoglobulin kappa light chain were synthesized in vitro in isolated mouse myeloma nuclei using bound endogenous RNA polymerase (RNA nucleotidyltransferase; nucleoside triphosphate:RNA nucleotidyltransferase; EC 2.7.7.6) and from isolated myeloma chromatin using exogenous Escherichia coli RNA polymerase. The in vitro RNA was transcribed using 5-mercuriuridine triphosphate and separated from in vivo RNA by chromatography on an agarose sulfhydryl affinity column. Template restriction is retained in vitro since synthesis of kappa chain messenger RNA, As determined by hybridization with complementary DNA, was much more efficient in nuclei and chromatin isolated from myeloma 66.2 tissue culture cells, a kappa-chain-producing cell line, than from MOPC 315 tissue culture cells, a lambda-chain-producing cell line. Transcription of kappa chain messenger RNA was 25 times more efficient in myeloma 66.2 nuclei than in myeloma 66.2 chromatin.