Present problems of somatic cell genetics]

A review of recent advances in genetics of somatic cells is given in the article. It contains three sections: 1) hybridization of somatic cells and its application to the mapping of genes and to the study of gene action; 2) relation of somatic cell genetics to genetic engineerings; 3) progress and perspectives of higher plant somatic cell genetics.     

Nuclear reprogramming in mammalian somatic cell nuclear cloning

Nuclear cloning is still a developing technique used to create genetically identical animals by somatic cell nuclear transfer into unfertilized eggs. Despite an intensive effort in a number of laboratories, the success rate of obtaining viable offspring from this technique remains less than 5%. In the past few years many investigators reported the reprogramming of specific nuclear activities in cloned animals, such as genome-wide gene expression patterns, DNA methylation, genetic imprinting, histone modifications and telomere length regulation. The results highlight the tremendous difficulty the clones face to reprogram the original differentiation status of the donor nuclei. Nevertheless, nuclei prepared from terminally differentiated lymphocytes can overcome this barrier and produce apparently normal mice. Study of this striking nuclear reprogramming activity should significantly contribute to our understanding of cell differentiation in more physiological settings.     

Identification of cells deficient in signaling-induced alternative splicing by use of somatic cell genetics

In recent years, a growing number of mammalian genes have been shown to undergo alternative splicing in response to extracellular stimuli. However, the factors and pathways involved in such signal-induced alternative splicing are almost entirely unknown. Here we describe a novel method for identifying candidate trans-acting factors that are involved in regulating mammalian alternative splicing, using the activation-induced alternative splicing of the human CD45 gene in T cells as a model system. We generated a cell line that stably expresses a CD45 minigene-based GFP reporter construct, such that the levels of green-fluorescent protein (GFP) expressed in the cell reflect the splicing state of the endogenous CD45 gene. Following mutagenesis of this cell line, and multiple rounds of selection for cells that displayed aberrant levels of GFP expression, we isolated several cell lines that are at least partially defective in their ability to support regulated alternative splicing of endogenous CD45 pre-mRNA in response to cell stimulation. Thus we have successfully isolated mutants in a mammalian alternative splicing pathway through use of a somatic cell-based genetic screen. This study clearly demonstrates the feasibility of using genetic screens to further our understanding of the regulation of mammalian splicing, particularly as it occurs in response to environmental cues.     

Production of mammalian somatic cell hybrids by means of polyethylene glycol treatment.

Polyethylene glycol (PEG) is known to promote fusion of plant protoplasts. Various adaptations of this treatment to mammalian, including human, cell cultures are reported here. PEG is very effective in producing hybrids capable of indefinite multiplication even in cases, such as early passage human skin fibroblasts and lymphocytes, known to be highly recalcitrant to other treatments.     

Genetic engineering of a mouse: Dr. Frank Ruddle and somatic cell genetics

Genetic engineering is the process of modifying an organism's genetic composition by adding foreign genes to produce desired traits or evaluate function. Dr. Jon W. Gordon and Sterling Professor Emeritus at Yale Dr. Frank H. Ruddle were pioneers in mammalian gene transfer research. Their research resulted in production of the first transgenic animals, which contained foreign DNA that was passed on to offspring. Transgenic mice have revolutionized biology, medicine, and biotechnology in the 21st century. In brief, this review revisits their creation of transgenic mice and discusses a few evolving applications of their transgenic technology used in biomedical research.     

Cytogenetic abnormalities in mammalian somatic cells

The different types of cytogenetic abnormalities are considered which are used in classic cytogenetics for the estimation of the levels of chromosome apparatus damages. The possible causes of cytogenetic anomalies and a number of methods of micronucleus tests are discussed. It was shown that the different levels of genetic material organization influence the realization of DNA defects into cytogenetic abnormality.     

Transmission genetics of the somatic hybridization process in Nicotiana

Summary Callus protoplasts of a Nicotiana tabacum chlorophyll-deficient mutant were fused with mesophyll protoplasts from one of following five sources: 4 cmsanalogs of tobacco bearing the cytoplasms of N. plumbaginifolia, N. suaveolens, N. repanda, and N. undulata, respectively, as well as wild species N. glauca. In another series of experiments, callus protoplasts from the chlorophyll-deficient genome Su/Su mutant of tobacco were fused with mesophyll protoplasts of the wild species N. glauca and those of a plastome chlorophyll-deficient tobacco mutant. The screening of hybrids consisted of visual identification followed by mechanical isolation and cloning of heteroplasmic fusion products in microdroplets of nutrient medium. Studies of regenerated plants included the analyses of gross morphology of plants, leaf and flower morphology, analysis of chromosome size and morphology and chromosome numbers, studies of multiple molecular forms of esterase and amylase, analysis of chloroplast DNA restriction patterns and analyses of chlorophyll-deficiency controlled by Su and P ^– genes. The study of progeny of 41 clones representing all species' combinations demonstrated that regenarants of most (63%) clones from intraspecific (for nuclear genes) combinations were cybrid forms, whereas in the case of the fusion N. tabacum + N. glauca, the true nuclear hybrids prevailed and the proportion of cybrids did not exceed 26%. Clones regenerating both hybrid and cybrid plants from the same fusion product were also found.