Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Transgene”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 235 records · Page 13Linked to original sources
PubMed2004

Efficiency of transgenic rat production is independent of transgene-construct and overnight embryo culture.

The aim of the present work was to study factors affecting the efficiency of transgenic technology in rats. We investigated the possible effects of pronuclear microinjection of buffer or different DNA-constructs on survival and development of rat zygotes in vitro and in vivo as well as the influence of overnight culture of these embryos before transfer into pseudopregnant foster mothers. The survival rate of zygotes and their development to the two-cell and morula stage was not affected by pronuclear microinjection with different DNA-constructs or buffer. However, the development to the blastocyst stage was impaired. Nevertheless, there was no difference in blastocyst development between zygotes injected with DNA-constructs or with buffer. Neither was there a difference in cell number in in vitro cultured blastocysts resulting from pronuclear microinjection of a transgene compared with non-injected controls. The survival rate to term was about 30% irrespective of whether microinjected embryos were transferred immediately after microinjection or after overnight culture in vitro. However, a reduction in the survival to term was observed for non-injected zygotes when they were developed in vitro to the two-cell stage before transfer to a pseudopregnant female. The percentage of transgenic rats that resulted from microinjected zygotes was similar in all groups regardless of the DNA-construct used (2.7-10.0%). In conclusion, the main detrimental factor in the microinjection of rat zygotes is the introduction of solution in the pronucleus. Overnight culture of zygotes between microinjection and oviduct transfer does not decrease the efficiency of transgenic rat generation.

Animals↗
PubMed2002

The methods to generate transgenic animals and to control transgene expression.

Transgenic animals have been used for years to study gene function and to create models for the study of human diseases. This approach has become still more justified after the complete sequencing of several genomes. Transgenic animals are ready to become industrial bioreactors for the preparation of pharmaceuticals in milk and probably in the future in egg white. Improvement of animal production by transgenesis is still in infancy. Despite its intensive use, animal transgenesis is still suffering from technical limitations. The generation of transgenics has recently become easier or possible for different species thanks to the use of transposons or retrovirus, to incubation of sperm which DNA followed by fertilization by intracellular sperm injection or not and to the use of the cloning technique using somatic cells in which genes have been added or inactivated. The Cre-LoxP system is more and more used to withdraw a given sequence from the genome or to target the integration of a foreign DNA. The tetracycline system has been improved and can more and more frequently be used to obtain faithful expression of transgenes. Several tools: RNA forming a triple helix with DNA, antisense RNA including double strand RNA inducing RNA interference and ribozymes, and also expression of proteins having a negative transdominant effect, are tentatively being improved to inhibit specifically the expression of host or viral genes.All these techniques are expected to offer experimenters new and more precise models to study gene function even in large animals. Improvement of breeding by transgenesis has become more plausible including through the precise allele replacement in farm animals.

Animals↗
PubMed1997

Analysis of transgene integration sites in transgenic pigs by fluorescence in situ hybridization.

The production of pigs transgenic for human decay accelerating factor (hDAF) as potential donors for clinical organ xenotransplantation was reported several years ago. For this purpose it is required that high levels of hDAF are expressed at relevant sites in transplantable organs. Currently, homozygous lines have been produced as well as lines from crosses between heterozygous animals from different founder lines, termed 'jigsaw' pigs. The purpose of the 'jigsaw' crosses is to combine the desirable hDAF protein expression patterns found in different founder lines. Initial selection of the 'jigsaw' pigs is based on the inheritance of the hDAF integration sites from both lines. Litters with potential homozygous transgenics and 'jigsaw' transgenics were analysed by fluorescence in situ hybridization (FISH) and slot blot analysis. Results show that both slot blot analysis and FISH are suitable to distinguish between pigs that are heterozygous and homozygous fir hDAF. However, FISH has the advantage of producing results more rapidly. For the identification of 'jigsaw' pigs FISH analysis was required since slot blot analysis lacked the required accuracy. On basis of these results, FISH analysis was made part of the routine screening programme for hDAF transgenic pigs.

Animals↗
PubMed1989

Isotype exclusion and transgene down-regulation in immunoglobulin-lambda transgenic mice.

A given B lymphocyte makes an antibody containing either kappa- or lambda-light chains, but not both. This isotype exclusion is effected at the level of the rearrangement of the immunoglobulin gene segments, although by an unknown mechanism. An attractive possibility is that, following productive rearrangement of one of the light-chain loci, the newly synthesized light-chain polypeptide inhibits DNA rearrangement for the other isotype. To test such feedback regulation, we have created transgenic mice carrying a rearranged lambda 1-gene. By contrast with the B cells in normal newborn mice which are mainly kappa+lambda-, the B cells in the newborn transgenic mice express lambda- but not kappa-chains. We propose that the synthesis of any light chain, be it kappa or lambda, that allows expression of IgM on the cell surface results in a cessation of all V-J joining. Interestingly, the limited light-chain repertoire of the transgenic mice does not persist and most adult B cells express endogenous kappa-rearrangements and down-regulate the transgene.

Animals↗
PubMed2005

Inheritance of a recessive transgene-associated character controlling albinism in transgenic bean (Phaseolus vulgaris L.).

We identified a transgenic line exhibiting albinism during our work to introduce genes through genetic engineering in dry bean (Phaseolus vulgaris). The transgenic mother plant (R0) presented a normal phenotype and generated albino and normal green plants in the first generation (R1). The segregation ratio of the albino character in the R1 and R2 generations fitted the expected ratio for a character controlled by a single recessive gene linked to a foreign gus gene, suggesting that albinism could be a consequence of insertional mutation caused by introduction of the exogenous gene. Analysis by electron microscope revealed that the albino cells possessed no chloroplasts and a greater number of mitochondria when compared to normal green plants. This transgenic bean line may be used in understanding the genetic control of chloroplast genesis, for acquiring additional knowledge of genomic structure or in physiological studies. This is the first described transgene-associated mutant bean plant.

Chloroplasts↗
PubMed1989

Expression of TL, H-2, and chimeric H-2/TL genes in transgenic mice: abnormal thymic differentiation and T-cell lymphomas in a TL transgenic strain.

To investigate the genetic regulation of TL expression, 12 transgenic mouse strains on a C3H (TL-nonexpressing) background have been derived: two Tg.Tlaa-3 strains with Tlaa-3 isolated from A-strain TL+ thymocytes, four Tg.T3b strains with T3b from a TL+ leukemia arising in a C57BL/6 (TL-) mouse, three Tg.Con.3 strains with an H-2Kb/T3b chimeric gene (construct 3,5'flanking region and exon 1 of H-2Kb and exons 2-6 of T3b), one Tg.Con.4 strain with a T3b/H-2Kb chimeric gene (construct 4, 5' flanking region and exon 1 of T3b and exons 2-8 of H-2Kb), and two Tg.H-2Kb strains with H-2Kb. Expression of the transgenes was determined by the presence of TL or H-2Kb products or transcripts. Both Tg.Tlaa-3 strains expressed high levels of TL antigen in thymus, indicating that (i) the 9.6-kilobase Tlaa-3 DNA fragment contains sufficient information for correct tissue-specific expression in thymocytes and (ii) TL- thymocytes of C3H provide conditions for the transcriptional activation of Tlaa-3. In contrast, neither the four Tg.T3b strains nor the Tg.Con.4 strain expressed transgenes, indicating that (i) T3b lacks elements necessary for TL expression in normal thymocytes and (ii) the corresponding endogenous TL genes of C3H mice also lack these elements. The pattern of TL expression in two of the three Tg.Con.3 strains was similar to that of H-2Kb expression, indicating that transcription of this H-2Kb/T3b chimeric gene was driven by the regulatory sequences of H-2Kb. The thymuses of mice derived from the Tg.Tlaa-3-1 strain were smaller than C3H thymuses, and the surface phenotype of Tg.Tlaa-3-1 thymocytes resembled thymocyte precursors (TL+L3T4-Lyt-2-Thy-1+H-2+). These mice developed a high incidence of lymphomas with the same thymocyte precursor phenotype. The study of TL transgenic strains should prove useful in defining the role of TL in normal and abnormal T-cell differentiation.

Animals↗
PubMed1996

Transgenic mice carrying a human mutant superoxide dismutase transgene develop neuronal cytoskeletal pathology resembling human amyotrophic lateral sclerosis lesions.

Mutations in the human Cu,Zn superoxide dismutase gene (SOD1) are found in 20% of kindreds with familial amyotrophic lateral sclerosis. Transgenic mice (line G1H) expressing a human SOD1 containing a mutation of Gly-93 --> Ala (G93A) develop a motor neuron disease similar to familial amyotrophic lateral sclerosis, but transgenic mice (line N1029) expressing a wild-type human SOD1 transgene do not. Because neurofilament (NF)-rich inclusions in spinal motor neurons are characteristic of amyotrophic lateral sclerosis, we asked whether mutant G1H and/or N1029 mice develop similar NF lesions. NF inclusions (i.e., spheroids, Lewy body-like inclusions) were first detected in spinal cord motor neurons of the G1H mice at 82 days of age about the time these mice first showed clinical evidence of disease. Other neuronal intermediate filament proteins (alpha-internexin, peripherin) also accumulated in these spheroids. The onset of accumulations of ubiquitin immunoreactivity in the G1H mice paralleled the emergence of vacuoles and NF-rich spheroids in neurons, but they did not colocalize exclusively with spheroids. In contrast, NF inclusions were not seen in the N1029 mice until they were 132 days old, and ubiquitin immunoreactivity was not increased in the N1029 mice even at 199 days of age. Astrocytosis in spinal cord was associated with a marked increase in glial fibrillary acidic protein immunoreactivity in the G1H mice, but not in the N1029 mice. Finally, comparative studies revealed a striking similarity between the cytoskeletal pathology in the G1H transgenic mice and in patients with amyotrophic lateral sclerosis. These findings link a specific SOD1 mutation with alterations in the neuronal cytoskeleton of patients with amyotrophic lateral sclerosis. Thus, neuronal cytoskeletal abnormalities may be implicated in the pathogenesis of human familial amyotrophic lateral sclerosis.

Aging↗
PubMed1998

Effects of 50 Hz magnetic fields on UV-induced skin tumourigenesis in ODC-transgenic and non-transgenic mice.

PURPOSE: To study the possible role of 50 Hz magnetic fields (MF) in UV-induced skin tumourigenesis using a sensitive animal model. MATERIALS AND METHODS: Transgenic mice (line K2) over-expressing the human ornithine decarboxylase (ODC) gene and their non-transgenic littermates were exposed for 10.5 months to UV-only or a combination of UV and a continuous (100 microT) or an intermittent MF with varying intensity (1.3-130 microT). RESULTS: Both MF exposure and transgenicity enhanced the onset rate of macroscopically detectable tumours, but the effect was statistically significant only for the MF exposure (p < 0.015). The number of animals bearing malignant tumours was low and similar in all exposure groups. Epidermal cysts (EC) appeared to be strongly associated with both MF exposure and high ODC activity (transgenic animals). However, EC are not known to be associated with carcinogenesis. The UV-only or combined UV and MF exposure did not affect the ODC activities measured at the end of the exposure. CONCLUSIONS: These results support the proposed tumour-promoting effect of MF, but do not suggest an important role for increased ODC activity in this process.

Animals↗
PubMed1994

Delayed morbidity and mortality of albumin/SV40 T-antigen transgenic mice after insertion of an alpha-fetoprotein/herpes virus thymidine kinase transgene and treatment with ganciclovir.

The alpha-fetoprotein (AFP) gene is normally expressed in fetal liver and transcriptionally silent in adult tissues, but can be abnormally reactivated in hepatocellular carcinoma (HCC). We linked 7.6 kb of 5'-flanking DNA from the mouse AFP gene to the herpes simplex virus (HSV) thymidine kinase gene (tk), and a line of transgenic mice was produced that expressed TK in a pattern similar to endogenous AFP. When these AFP/tk transgenic mice were crossed to another transgenic line that develops multifocal HCC due to expression of a SV40 large T-antigen transgene under regulation of the albumin promoter/enhancer complex, a significant delay of tumor progression could be achieved by administration of ganciclovir (GCV), a cytotoxic compound that is a substrate for phosphorylation by viral, but not mammalian, TK. Control animals carrying only the tk gene were unaffected by GCV treatment. These results illustrate the feasibility of prophylactic gene therapy for ablation of cancer, utilizing a strategy in which the tk gene is regulated by a promoter expected to be active only in tumor cells.

Albumins↗
PubMed1996

Formation and persistence of O6-ethylguanine in genomic and transgene DNA in liver and brain of lambda(lacZ) transgenic mice treated with N-ethyl-N-nitrosourea.

LacZ transgenic mice are suitable for short-term mutagenicity studies in vivo. Mutagenicity in these mice is determined in the lacZ transgene. Since the lacZ gene is of bacterial origin the question has been raised whether DNA-adduct formation and repair in the transgene are comparable to those in total genomic DNA. Mice were treated with N-ethyl-N-nitrosourea (ENU) and killed at several time points following treatment. Some mice were pretreated with O6-benzylguanine to inactivate the repair protein O6-alkylguanine-DNA alkyltransferase (AGT). O6-ethylguanine (O6-EtG) was determined in lacZ in liver and brain by means of a monoclonal antibody-based immunoaffinity assay. In addition, O6-EtG and N7-ethylguanine (N7-EtG) were assayed in total genomic DNA of liver and brain with an immunoslotblot procedure. In liver, the initial O6-EtG level in total genomic DNA was 1.6 times that in lacZ. The extent of repair of O6-EtG during the first 1.5 h after treatment was 2.1 times that in lacZ. At later time points, O6-EtG repair was the same. N7-EtG repair in genomic DNA was evident. In contrast to the liver, little repair of O6-EtG in total genomic and lacZ DNA occurred in the brain while N7-EtG was repaired. No initial difference in O6-EtG levels were found in lacZ and genomic brain DNA. These findings indicate that in the liver, total genomic DNA is more accessible than lacZ to ENU and/or the AGT protein, during the first 1.5 h following treatment. Because the difference in O6-EtG levels in the transgene and genomic DNA in the liver is restricted to the first 1.5 h after treatment, while the fixation of mutations occurs at later time points, O6-EtG-induced mutagenesis most likely is also very similar in both types of DNA.

Animals↗
PubMed1989

A salivary amylase transgene is efficiently expressed in liver but not in parotid gland of transgenic mice.

Two distinct mouse amylase cDNAs, corresponding to the genes Amy-1.1 and Amy-1.2, have been isolated from a YBR/Ki parotid cDNA library. A cosmid clone containing the intact Amy-1.1 gene from strain YBR/Ki, including both the parotid and liver promoters, was transferred to the germ line of C57BL/6J mice. Two independent transgenic lines were produced. The transferred genes are organized as a 4-copy autosomal locus in line Tg518 and an 8-copy Y-linked locus in line Tg2736. Serum of both transgenic lines contained high levels of the AMY1B isozyme encoded by the transferred gene. Transcripts were detected in liver and, at a lower level, in several other tissues including white and brown fat. The anticipated expression in parotid was not observed. Constructs containing 270 or 540 bp of the 5' flanking region of the parotid promoter, cloned upstream of the chloramphenicol acetyltransferase (CAT) structural gene, were also not expressed in transgenic mice. The results suggest that sequences located more than 5 kb upstream of the Amy-1 parotid promoter and/or more than 10 kb downstream from the structural gene are required for parotid-specific expression. The results also demonstrate that non-parotid sources can produce a normal level of AMY1 in serum. Liver is the probable source of AMY1 in serum of these transgenic mice.

Amylases↗
PubMed2005

Expression of complementary RNA from chloroplast transgenes affects editing efficiency of transgene and endogenous chloroplast transcripts.

The expression of angiosperm chloroplast genes is modified by C-to-U RNA editing. The mechanism for recognition of the approximately 30 C targets of editing is not understood. There is no single consensus sequence surrounding editing sites, though sites can be grouped into small 'clusters' of two to five sites exhibiting some sequence similarity. While complementary RNA that guides nucleotides for alteration has been detected in other RNA modification systems, it is not known whether complementary RNA is involved in chloroplast editing site recognition. We investigated the effect of expressing RNA antisense to the sequences -20 to +6 surrounding the RpoB-2 C target of editing, which is a member of a cluster that includes the PsbL-1 and Rps14-1 sites. Previous experiments had shown that chloroplast rpoB transgene transcripts carrying only these 27 nt were edited in vivo at the proper C. Though transcripts carrying sequences -31 to +60 surrounding the RpoB-2 sites were edited in chloroplast transgenic plants, transcripts carrying the -31 to +62 region followed by the 27 nt complementary region were not edited at all. In contrast, a similar construct, in which the C target as well as the preceding and subsequent nucleotides were mismatched within the 27 nt region, was efficiently edited. The presence of any of the four transgenes carrying RpoB-2 sequences in sense and/or antisense orientation resulted in reduced editing at the PsbL-1 site. Chloroplast transgenic plants expressing the three different antisense RNA constructs exhibited abnormal growth and development, though plants expressing the 92 nt sense transcripts were phenotypically normal.

Chloroplasts↗
PubMed2005

Low frequency of transgene flow from Bt/CpTI rice to its non-transgenic counterparts planted at close spacing.

Crop-to-crop transgene flow will affect seed purity of non-GM rice varieties, leading to unwanted consequences. To assess the maximum probability of transgene outflow in rice (Oryza sativa), gene flow experiments were conducted with three cultivation patterns with different mixed-planting proportions of adjacent GM and non-GM rice at two sites in Fujian and Hainan Provinces of China. Three GM rice lines containing two insect-resistance genes (Bt/CpTI) and their non-GM counterparts were used in the experiments to allow natural hybridization to occur. A hygromycin resistance gene was used as a selective marker for identifying hybrids. Based on the examination of > 645 700 geminated seeds, the result showed low frequencies (0.05-0.79%) of transgene flow from GM to non-GM rice at close spacing, although with significant variation among mixed-planting proportions. It is concluded that rice transgene flow will occur at a very low frequency (< 1.0%), even if the GM rice is planted at close spacing with non-GM rice, and high densities of GM rice cultivated in the neighborhood of non-GM rice will increase the probability of outcrossing with the non-GM rice.

China↗
PubMed2004

Differences of globin transgene expression in stably transfected cell lines and transgenic mice.

Previous studies demonstrated that DNase I hypersensitive site -40 (HS-40) of the alpha-globin locus is capable of greatly enhancing expression of a hybrid beta/gamma-globin transcriptional unit in plasmid-transfected murine erythroleukemia (MEL) cells. However, as reported here, this same gamma-globin gene expression cassette was only transcribed at trace amounts in erythroid cells of transgenic mice. This lack of expression was not directly attributable to the beta/gamma-globin transcriptional unit, since this same unit linked to a composite beta-globin locus control region was expressed at high levels in transgenic mice. This lack of expression was also not directly attributable to chromosomal position effects, since addition of chromatin insulators failed to increase the frequency of expression. DNase I hypersensitivity and chromatin immunoprecipitation assays demonstrated that the lack of expression was correlated with a closed chromatin structure. We hypothesize that transgenes undergo dynamic changes in chromatin conformation following chromosomal integration and that the discrepant results reported here can be attributed to the relatively high level of chromatin remodeling that occurs in the transgenic mouse model, coupled with the relative inability of the HS-40 element to maintain an open chromatin state under such conditions.

Animals↗
PubMed1994

Effects of glucose on insulin secretion, glucokinase activity, and transgene expression in transgenic mouse islets containing an upstream glucokinase promoter-human growth hormone fusion gene.

We have analyzed in organ culture the effects of glucose on glucose-induced insulin secretion, glucokinase (GK) activity, and human growth hormone (hGH) expression in pancreatic islets from transgenic mice containing an upstream GK promoter-hGH fusion gene. Freshly isolated islets from these mice had a normal insulin secretory response to glucose but showed subtle defects after culture in low or high glucose for 4 days that may have been due to the accumulation of hGH in the culture media. Islets cultured from both normal and transgenic mice had approximately a fourfold induction of GK activity in response to an increased concentration of glucose in the culture media, whereas no such change in total islet hGH production was observed. Immunocytochemical localization of hGH in islets cultured in 3 mM glucose showed a pattern similar to that in freshly isolated islets. However, after culture in 30 mM glucose, hGH immunostaining became strikingly more heterogeneous. We conclude 1) that GK-hGH transgene expression does not appear to adversely affect glucose-stimulated insulin secretion in vivo or in freshly isolated islets, 2) that glucose does not induce transgene expression, thus providing additional evidence against an effect of glucose on GK gene transcription in the islet, and 3) that glucose stimulates the co-release of hGH with insulin, thereby enhancing the heterogeneous staining pattern seen among pancreatic beta-cells.

Animals↗
PubMed1989

Lymphomagenesis in Emu-myc transgenic mice does not require transgene rearrangement or mutation of myc exon 1.

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.

Amino Acid Sequence↗
PubMed1995

Ig gamma 2b transgenes promote B cell development but alternate developmental pathways appear to function in different transgenic lines.

Analysis of B cell development in three strains of gamma 2b transgenic mice shows that the gamma 2b H chain can replace the microH chain in promoting B cell differentiation. The 348C line produces 90% gamma 2b-only B cells and 10% B cells; which co-express gamma 2b and endogenous sIgM and sIgD. These IgG2b+ B cells develop into mature, recirculating CD23+ B cells. The 343-1 and gamma 2b-T15 transgenic mice produce sIgMhigh:sIgDlow:CD23- B cells that generally co-express the gamma 2b transgene-encoded H chain. Such B cells are either developmentally arrested immature B cells or arise from B-1 (CD5) progenitors. The gamma 2b-T15 mice can produce gamma 2b-only CD23+ B cells following inactivation of the endogenous mu locus, whereas 343-1 mice fail to develop B cells. Thus, gamma 2b H chains: 1) can act alone to promote the development of mature B cells, 2) synergize with microH chains for allelic exclusion, and 3) vary in their influence on B cell development in different transgenic mouse strains.

Animals↗
PubMed1995

A transgenic mouse model (TG.AC) for skin carcinogenesis: inducible transgene expression as a second critical event.

The v-Ha-ras transgenic TG.AC mouse line behaves as a genetically initiated model for mouse skin tumorigenesis with enhanced susceptibility to skin carcinogens. TG.AC mice develop epidermal papillomas in fewer than 20 weeks in response to the topical application of a variety of chemicals such as complete carcinogens, phorbol ester-type tumor promoters, and nonphorbol ester-type tumor promoters as well as to full-thickness skin wounds or plucking of the dorsal hair. We have found that the pedunculated epidermal papillomas can arise as focal hyperplasias from the permanent portion of the follicular epithelium. Expression of the v-Ha-ras transgene serves as a marker for tumor development since it is expressed at significant levels in the papilloma precursors, focal follicular hyperplasias, and the papillomas but not in the surrounding skin. Transgene expression colocalizes with increased cell proliferation in the papillomas as compared to non-tumor bearing surrounding skin. Malignant skin tumors, primarily squamous cell carcinomas and sarcomas, develop from sites of papilloma development in approximately 40% of papilloma bearing mice. As well as significant levels of transgenic v-Ha-ras expression, some of the malignancies also exhibit karyotypic changes which include trisomy of chromosome six or fifteen, but not chromosome seven. We believe that the TG.AC mouse line serves not only as a model for studying the mechanisms of skin tumorigenesis, but will also be a useful adjunct to the two year NTP toxicity/carcinogenicity studies by identifying carcinogens in fewer than 20 weeks.

Animals↗