Molecular biology & medicine最新文献

In most human Burkitt's lymphomas, translocation of the myc oncogene to an immunoglobulin locus is associated with loss of myc exon 1 or with mutations near its 3' border, a region where myc transcription is attenuated and translation of a larger myc polypeptide initiates. Emu-myc transgenic mice, which bear the three myc exons coupled to an immunoglobulin enhancer, provide a model for the development of such lymphomas, because their lymphomagenesis appears to require events other than expression of the transgene. To determine whether myc rearrangement or exon 1 mutation is a necessary tumorigenic event, we examined the transgene structure and myc exon 1 sequences in Emu-myc B lymphoid tumours. Southern blots revealed no transgene rearrangements in 20 of the lymphomas, and only two tumours showed amplification (2 to 5-fold). To search for exon 1 alterations, the exon 1 mRNA region was amplified from five tumours by polymerase chain reaction and sequenced, but no mutations were found. Hence, neither excision nor mutation of exon 1 is necessary to render myc tumorigenic. The sequence analysis across the exon 1-exon 2 boundary unexpectedly revealed an ambiguity in myc splicing that predicts a variant form of the larger myc polypeptide lacking a single amino acid residue.

Genetically altered embryonic stem (ES) cells re-injected into mouse blastocysts take part in the formation of all tissues, including the germ line, thus generating transgenic offspring. This approach in combination with the homologous recombination technology offers the possibility of altering ES cells in a controlled manner and therefore of generating transgenic mice with a predetermined genome. We summarize here advances in mouse embryology and genetics that have led to this exciting development.

Recent developments in molecular biology technology have greatly facilitated the methods for detection of mutations responsible for genetic disorders in both humans and animals. In this article we review some of these new methods and present a diagnostic algorithm that facilitates the routine and rapid diagnosis of any genetic disorder for which the defective gene has been isolated.

The study of mammalian development has very quickly moved from a largely descriptive endeavour to one in which very precise mechanistic questions can now be formulated and answered. Undoubtedly, advances in this area have been the result of a strong foundation in experimental embryology, the application of molecular genetic techniques to the isolation and analysis of genes of developmental interest, and the ability to manipulate genetically the embryo through transgenic mouse technology. Perhaps the most dramatic illustration of the power of these new technologies is the potential ability to generate mice either that carry mutations in virtually any gene in the germ line through gene targeting in totipotent embryonic stem (ES) cells or that lack specific cell types through the genetic ablation technology reviewed here. Together, these two approaches have made it possible to knock out either a specific gene or a specific cell type in an intact animal and thus offer almost unlimited possibilities for addressing questions concerning the molecular and cellular biology of development. As well, animal models for various human diseases such as dwarfism, immunodeficiencies and demyelination can now be generated. It is clear that further refinements in both gene targeting and genetic ablation technologies are necessary before the full potential of either approach will be realized. Further development of conditional or inducible ablation strategies, coupled with a more precise definition of the cis-acting sequences, responsible for directing gene expression in fully differentiated and more primitive cells, will greatly broaden the range of questions that can be addressed by this approach.(ABSTRACT TRUNCATED AT 250 WORDS)

To investigate the function of activated oncogenes we attempted to create transgenic mice carrying activated human c-Ha-ras genes which have their own promoters. However, we never obtained any transgenic pups which developed to term, because all transgenic embryos were malformed, became developmentally arrested conceptuses or developed embryonic tumors during ontogenesis. The mRNA expression of the transgenes was detected in two tumors obtained after introduction of the DNA fragment containing the activated human c-Ha-ras gene for p21 with valine at the 12th codon or with leucine at the 61st codon. Histological analysis indicated that each tumor consisted of at least three types of cells: two originating from different germ layers (the endoderm in one case and the mesoderm in the other) and the third from extra embryonic ectoderm. It was suggested that the activated human c-Ha-ras gene has a critical effect on the development of tumors in normal embryos as well as in transformation of NIH3T3 cells.

Transgenic mice harbouring growth hormone gene constructs have been produced by DNA microinjection into pronuclei of fertilized oocytes. We examined transgenic mice carrying a mouse metallothionein I-human growth hormone (mMT I-hGH) fusion gene. Here, we present our results concerning gene integration, gene expression, and phenotypical, clinical and pathomorphological alterations found in mice expressing the hGH transgene. Body and organ growth was significantly increased in transgenic mice, whereas fertility was found to be reduced. The life-span was markedly shortened indicating detrimental side-effects of the high levels of circulating hGH. Lesions of kidneys, liver and heart were the predominant pathological findings. Our own results are compared with those obtained by other authors who have investigated mice carrying rat, bovine or ovine growth hormone fusion genes. GH-transgenic mice may serve as a model system to investigate ectopic expression of hormone genes thus circumventing endogenous feedback control mechanisms in complex hormonal cascades.

The development of a new mammalian genetics in which specifically designed gene alterations of pre-existing endogenous genetic loci may be achieved via tissue culture of stem cells is described. At present this technology is rapidly emerging for mice by gene targeting in cultured embryonic stem cells, isolation of the desired cell clone and re-constitution of germ-line chimaeric animals. Results to date and strategies for gene targeting and isolation of the required cell clones are discussed. This new genetic technology is likely to have a major impact both in genetic studies and, especially if it can be extended to larger mammalian species, in practical applications.