Malcolm L. Gefter

A simple method for polyethylene glycol-promoted hybridization of mouse myeloma cells.

A simple method is described for promoting the fusion of mouse myeloma cells in suspension with polyethylene glycol (PEG 1000). By carefully controlling the concentration of PEG and the time of exposure of the cells, it was possible to obtain hybridization frequencies several-hundred-fold higher than those obtained with Sendai virus.

DNA synthesis in cell-free extracts of a DNA polymerase-defective mutant

Abstract A DNA-synthesizing activity has been isolated from an E. coli mutant defective in DNA polymerase. Like DNA polymerase, the system requires the presence of all four deoxynucleoside triphosphates, magnesium ion and a native DNA template for maximal activity. The activity can be distinguished from E. coli DNA polymerase on the basis of its sensitivity to high ionic strength and to p-chloromercuribenzoate. The activity is insensitive to antiserum directed against E. coli DNA polymerase. Our results do not exclude the possibility that the activity isolated is composed of, in part, an altered form of DNA polymerase.

Bacteriophage φ80-induced low molecular weight RNA

Abstract A species of low molecular weight RNA (M 3 ) which is produced in bacteriophage φ80-infected Escherichia coli has been isolated and characterized. M 3 appears immediately after infection and its synthesis continues until lysis. This RNA is unstable; its mean half-life is 13.5 min. The structural gene for M 3 is localized on the φ80 genome to the right of the exonuclease locus and to the left of gene Q. The complete nucleotide sequence of M 3 has been determined. It is 62 nucleotides in length. The 5′ end begins with pppGp and the 3′ terminus is C-U-U-U-U-U-A-A OH . The sequence and sensitivity to ribonuclease digestion suggests a highly double-stranded structure for M 3 .

Regulation of Immunoglobulin Expression in Mouse Myeloma Cells

Recent experimental evidence suggests that all the somatic cells of a given individual have considerable genetic potential and may be genetically identical (Gurdon 1974; Illmensee and Mintz 1976). In addition, the vertebrate cell contains a number of genes which encode the constant regions of the different classes and subclasses of immunoglobulin heavy (H) and light (L) polypeptide chains, as well as the genes for some, if not all, of the heavyand light-chain variable regions that the individual can express. Furthermore, the cells of a heterozygous individual contain the information to synthesize two different allelic products of each gene (Cebra 1969). In spite of this large genetic potential, most somatic cells never express any immunoglobulin (Ig) genes at all, and the fully differentiated plasma cell usually produces large amounts of only one class or subclass of H and L chain and one set of variable regions. The molecular mechanisms responsible for this kind of restricted expression are not known. Some of these controls may be the same as those which regulate the expression of genes other than those for immunoglobulin or which govern phenomena such as X-chromosome inactivation (Lyon 1972). However, there is reason to believe that there are also mechanisms that are either unique to immunoglobulin expression or are at least restricted to a limited set of traits. The molecular events that (a) permit two (or perhaps more [Capra and Kindt 1975]) genes to code for one polypeptide chain, (b) constrain individual cells of heterozygous animals to express only one of two alleles (Cebra 1969), (c) direct the coordinated expression of particular H-L gene pairs, or (d) function in the generation of antibody diversity may represent such unique mechanisms. In an attempt to learn more about these mechanisms which regulate immunoglobulin expression, we have used genetic and biochemical tools to study cloned lines of mouse myeloma cells. In this paper, we wilt review the genetic information we have obtained by (1) isolating somatic cell hybrids of mouse myeloma cells expressing different classes and subclasses of immunoglobulin, and (2) isolating