Estimation of light-chain gene reiteration of mouse immunoglobulin by DNA-RNA hybridization.

A 12S species of RNA has been isolated from membrane-bound polyribosomes of MPC-11 mouse myeloma cells. This 12S RNA is translated by an extract of Krebs-II mouse ascites cells into immunoglobulin light chain. Labeled 12S RNA has been prepared by incubation of myeloma cells with [(3)H]uridine in the presence of low concentrations of actinomycin D and ethidium bromide. This RNA has been hybridized under conditions of DNA excess to mouse myeloma DNA and to liver DNA. A C(0)t(1/2) of about 150 has been obtained, corresponding to a reiteration frequency of about 40. Unlabeled 12S RNA competes with the labeled species in hybridization experiments, whereas globin mRNA or mouse ascites 12S RNA does not. It is suggested that 12S RNA hybridizes only to V(kappa)-genes of the same subclass, and that there may be several hundred genes coding for V(kappa)-regions of all subclasses in a mouse genome. Moreover, gene amplification in myeloma cells is not detected.