15 research outputs found

    Reduction of Werner Syndrome Protein Enhances G:C → A:T Transition by<i>O</i><sup>6</sup>‑Methylguanine in Human Cells

    No full text
    <i>O</i><sup>6</sup>-Methylguanine (<i>O</i><sup>6</sup>-MeG) is a damaged base produced by methylating reagents. The Werner syndrome protein (WRN) is a cancer-related human DNA helicase. The effects of WRN reduction on <i>O</i><sup>6</sup>-MeG-caused mutagenesis were assessed by an siRNA-mediated knockdown in human U2OS cells, using a shuttle plasmid with a single <i>O</i><sup>6</sup>-MeG base in the <i>supF</i> gene. The plasmid DNA was replicated in the cells, isolated, and electroporated into an <i>Escherichia coli</i> indicator strain. The lowered amount of WRN increased the frequency of mutations induced by <i>O</i><sup>6</sup>-MeG, mainly G:C → A:T substitution. The increased mutation rate suggested that the cancer-related WRN suppresses the G:C → A:T substitution by <i>O</i><sup>6</sup>-MeG in human cells

    Strategy for re-expression of MSH2 from the endogenous promoter by gene targeting.

    No full text
    <p>The MSH2 gene is located on chromosome 2 and the indicated allele has the region from exon 1 to exon 8 of MSH2. Region from exon 9 to 16 is deleted in both chromosomes. The synthetic exon from exon 9 to exon 16 was introduced into downstream of exon 8 by targeting. After Cre recombination, Neo<sup>R</sup> is removed, and lox71 and lox66 creates a defective lox sequence, which is no longer a target of Cre recombinase. DT-A stands for diphtheria toxin-A.</p

    Establishment of Nalm-6-MSH+.

    No full text
    <p>The MSH2-expressing cell lime, i.e., Nalm-6-MSH+, has been established by introduction of cDNA of exon 9 to 16 of the MSH2 gene into the original Nalm-6 cells. The resulting cell line exhibits high efficiency of gene targeting as the original Nalm-6 and is genetically stable. It is also resistant to killing effects of alkylating agents.</p

    The CGH array analysis of the MSH2 gene in Nalm-6 genome.

    No full text
    <p>Part of the chromosome 2 covering the MSH2 gene is presented. The chromosome region where one allele is deleted is boxed with black color and the region where both alleles are missed is boxed with red color. The region covering MSH2 is indicated with a red thick line.</p

    Gene targeting vectors for knockout and knock-in of the REV3L gene.

    No full text
    <p>The exon 5 is replaced with the hygromycin-resistance gene for knockout (A). The mutations that direct substitution of amino acids in the catalytic site of REV3L, i.e., D2781A and D2783A, were introduced into exon 30 for knock-in (B).</p

    Gene targeting efficiency at REV3 loci using targeting vectors for the knockout mutation (knockout) and the catalytically dead mutation (knock-in).

    No full text
    <p>Gene targeting efficiency at REV3 loci using targeting vectors for the knockout mutation (knockout) and the catalytically dead mutation (knock-in).</p

    Crystal Structure and Cyclic Hydrogenation Property of Pr<sub>4</sub>MgNi<sub>19</sub>

    No full text
    The hydrogen absorption–desorption property and the crystal structure of Pr<sub>4</sub>MgNi<sub>19</sub> was investigated by pressure–composition isotherm measurement and X-ray diffraction (XRD). Pr<sub>4</sub>MgNi<sub>19</sub> consisted of two phases: 52.9% Ce<sub>5</sub>Co<sub>19</sub>-type structure (3R) and 47.0% Gd<sub>2</sub>Co<sub>7</sub>-type structure (3R). Sm<sub>5</sub>Co<sub>19</sub>-type structure (2H) and Ce<sub>2</sub>Ni<sub>7</sub>-type structure (2H) were not observed in the XRD profile. The Mg atoms substituted at the Pr sites in a MgZn<sub>2</sub>-type cell. The maximum hydrogen capacity reached 1.14 H/M (1.6 mass%) at 2 MPa. The hysteresis factor, Hf = ln­(<i>P</i><sub>abs</sub>/<i>P</i><sub>des</sub>), was 1.50. The cyclic hydrogenation property of Pr<sub>4</sub>MgNi<sub>19</sub> was investigated up to 1000 absorption–desorption cycles. After 250, 500, 750, and 1000 cycles, the retention rates of hydrogen were reduced to 94%, 92%, 91%, and 90%, respectively. These properties were superior to those of Pr<sub>2</sub>MgNi<sub>9</sub> and Pr<sub>3</sub>MgNi<sub>14</sub>

    Gene targeting efficiency at <i>HPRT</i> loci using targeting vectors without mismatch sequences with mismatch sequences.

    No full text
    <p>Gene targeting efficiency at <i>HPRT</i> loci using targeting vectors without mismatch sequences with mismatch sequences.</p

    Crystal Structure and Cyclic Hydrogenation Property of Pr<sub>4</sub>MgNi<sub>19</sub>

    No full text
    The hydrogen absorption–desorption property and the crystal structure of Pr<sub>4</sub>MgNi<sub>19</sub> was investigated by pressure–composition isotherm measurement and X-ray diffraction (XRD). Pr<sub>4</sub>MgNi<sub>19</sub> consisted of two phases: 52.9% Ce<sub>5</sub>Co<sub>19</sub>-type structure (3R) and 47.0% Gd<sub>2</sub>Co<sub>7</sub>-type structure (3R). Sm<sub>5</sub>Co<sub>19</sub>-type structure (2H) and Ce<sub>2</sub>Ni<sub>7</sub>-type structure (2H) were not observed in the XRD profile. The Mg atoms substituted at the Pr sites in a MgZn<sub>2</sub>-type cell. The maximum hydrogen capacity reached 1.14 H/M (1.6 mass%) at 2 MPa. The hysteresis factor, Hf = ln­(<i>P</i><sub>abs</sub>/<i>P</i><sub>des</sub>), was 1.50. The cyclic hydrogenation property of Pr<sub>4</sub>MgNi<sub>19</sub> was investigated up to 1000 absorption–desorption cycles. After 250, 500, 750, and 1000 cycles, the retention rates of hydrogen were reduced to 94%, 92%, 91%, and 90%, respectively. These properties were superior to those of Pr<sub>2</sub>MgNi<sub>9</sub> and Pr<sub>3</sub>MgNi<sub>14</sub>

    Comparison of Nalm-6 (MSH-) and Nalm-6-MSH+.

    No full text
    <p>Western blot analysis of MSH2/MSH6 expression (A) and RT-PCR analysis of MSH2 mRNA from exon 8 to 16 (B).</p
    corecore