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Gene Trapping (gene + trapping)

Distribution by Scientific Domains

Life Sciences	85%

Selected Abstracts

Sorting nexin-14, a gene expressed in motoneurons trapped by an in vitro preselection method

DEVELOPMENTAL DYNAMICS, Issue 4 2001
Patrick Carroll
Abstract A gene-trap strategy was set up in embryonic stem (ES) cells with the aim of trapping genes expressed in restricted neuronal lineages. The vector used trap genes irrespective of their activity in undifferentiated totipotent ES cells. Clones were subjected individually to differentiation in a system in which ES cells differentiated into neurons. Two ES clones in which the trapped gene was expressed in ES-derived neurons were studied in detail. The corresponding cDNAs were cloned, sequenced, and analysed by in situ hybridisation on wild-type embryo sections. Both genes are expressed in the nervous system. One gene, YR-23, encodes a large intracellular protein of unknown function. The second clone, YR-14, represents a sorting nexin (SNX14) gene whose expression in vivo coincides with that of LIM-homeodomain Islet-1 in several tissues. Sorting nexins are proteins associated with the endoplasmic reticulum (ER) and may play a role in receptor trafficking. Gene trapping followed by screening based on in vitro preselection of differentiated ES recombinant clones, therefore, has the potential to identify integration events in subsets of genes before generation of mouse mutants. © 2001 Wiley-Liss, Inc. [source]

A review of current large-scale mouse knockout efforts

GENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 2 2010
Chunmei Guan
Abstract After the successful completion of the human genome project (HGP), biological research in the postgenome era urgently needs an efficient approach for functional analysis of genes. Utilization of knockout mouse models has been powerful for elucidating the function of genes as well as finding new therapeutic interventions for human diseases. Gene trapping and gene targeting are two independent techniques for making knockout mice from embryonic stem (ES) cells. Gene trapping is high-throughput, random, and sequence-tagged while gene targeting enables the knockout of specific genes. It has been about 20 years since the first gene targeting and gene trapping mice were generated. In recent years, new tools have emerged for both gene targeting and gene trapping, and organizations have been formed to knock out genes in the mouse genome using either of the two methods. The knockout mouse project (KOMP) and the international gene trap consortium (IGTC) were initiated to create convenient resources for scientific research worldwide and knock out all the mouse genes. Organizers of KOMP regard it as important as the HGP. Gene targeting methods have changed from conventional gene targeting to high-throughput conditional gene targeting. The combined advantages of trapping and targeting elements are improving the gene trapping spectrum and gene targeting efficiency. As a newly-developed insertional mutation system, transposons have some advantages over retrovirus in trapping genes. Emergence of the international knockout mouse consortium (IKMP) is the beginning of a global collaboration to systematically knock out all the genes in the mouse genome for functional genomic research. genesis 48:73,85, 2010. © 2010 Wiley-Liss, Inc. [source]

Characterizing the regulation of the Pu promoter in Acinetobacter baylyi ADP1

ENVIRONMENTAL MICROBIOLOGY, Issue 7 2008
Wei E. Huang
Summary Effective gene trapping and screening requires sensory and regulatory compatibility of both host and exogenous systems. The naturally competent bacterium Acinetobacter baylyi ADP1 is able to efficiently take up and integrate exogenous DNA into the chromosome, making it an attractive host system for a wide range of metagenomic applications. To test the ability of A. baylyi ADP1 to express the XylR-regulated Pu promoter from Pseudomonas putida mt-2, we have constructed and examined an A. baylyi ADP1 strain, ADPWH- Pu-lux-xylR. The Pu promoter in ADPWH- Pu-lux-xylR was specifically induced by toluene, m -, p - and o- xylene. The substrate-induced Pu promoter was highly dependent on the growth medium: it was repressed in rich media until stationary phase, but was immediately induced in minimal medium with glucose as the sole carbon source (MMG). However, the Pu promoter was repressed in MMG when it was supplemented with 5 g l,1 yeast extract. Further investigation showed that the Pu promoter in MMG was repressed by 0.5 g l,1 aspartic acid or asparagine, but not repressed by glutamine. Changing the carbon/nitrogen ratios by addition of ammonia did not significantly affect the Pu promoter activity but addition of nitrate did. These results show that A. baylyi ADP1 reproduced characteristics of the XylR-regulated Pu promoter observed in its original host. It demonstrates that A. baylyi could provide an excellent genetic host for a wide range of functional metagenomic applications. [source]

A review of current large-scale mouse knockout efforts

A bidirectional gene trap construct suitable for T-DNA and Ds -mediated insertional mutagenesis in rice (Oryza sativa L.)

PLANT BIOTECHNOLOGY JOURNAL, Issue 5 2004
Andrew L. Eamens
Summary A construct suitable for genome-wide transfer-DNA (T-DNA) and subsequent transposon-based (Ds) gene trapping has been developed for use in rice (Oryza sativa). This T-DNA/Ds construct contains: Ds terminal sequences immediately inside T-DNA borders for subsequent Ds mobilization; promoterless green fluorescent protein (sgfpS65T) and ,-glucuronidase (uidA) reporter genes, each fused to an intron (from Arabidopsis GPA1 gene) to enable bidirectional gene trapping by T-DNA or Ds; an ampicillin resistance gene (bla) and a bacterial origin of replication (ori) to serve as the plasmid rescue system; an intron-containing hygromycin phosphotransferase gene (hph) as a selectable marker or Ds tracer; and an intron-containing barnase gene in the binary vector backbone (VB) to select against transformants carrying unwanted VB sequences. More than a threefold increase over previously reported reporter gene-based gene trapping efficiencies was observed in primary T-DNA/Ds transformant rice lines, returning an overall reporter gene expression frequency of 23%. Of the plant organs tested, 3.3,7.4% expressed either reporter at varying degrees of organ or tissue specificity. Approximately 70% of the right border (RB) flanking sequence tags (FSTs) retained 1,6 bp of the RB repeat and 30% of the left border (LB) FSTs retained 5,23 bp of the LB repeat. The remaining FSTs carried deletions of 2,84 bp inside the RB or 1,97 bp inside the LB. Transposition of Ds from the original T-DNA was evident in T-DNA/Ds callus lines super-transformed with a transposase gene (Ac) construct, as indicated by gene trap reporter activity and rescue of new FSTs in the resulting double transformant lines. [source]

Gene trap mutagenesis in mice: New perspectives and tools in cancer research

CANCER SCIENCE, Issue 1 2008
Ken-ichi Yamamura
The complete human DNA sequence of the human genome was published in 2004 and we entered the postgenomic era. However, many studies showed that gene function is much more complex than we expected, and that mutation of disease genes does not give any clue for molecular mechanisms for disease development. Since the first report on gene knockout mice in 1989, knockout mice have been shown to be a powerful tool for functional genomics and for the dissection of developmental processes in human diseases. In accordance with this successful application of knockout mice, three major mouse knockout programs are now underway worldwide, to mutate all protein-encoding genes in mouse embryonic stem cells using a combination of gene trapping and gene targeting. We developed the exchangeable gene trap method suitable for large scale mutagenesis in mice. In this method we can produce null mutation and post-insertional modification, enabling replacement of the marker gene with a gene of interest and conditional knockout. We herein discuss the effect of this gene-driven type approach for cancer research, especially for finding the genes that are related to cancer, but are paid little attention in hypothesis-driven cancer research. (Cancer Sci 2008; 99: 1,6) [source]