In vivo genome editing restores haemostasis in a mouse model of haemophilia

Editing of the human genome to correct disease-causing mutations is a promising approach for the treatment of genetic disorders. Genome editing improves on simple gene-replacement strategies by effecting in situ correction of a mutant gene, thus restoring normal gene function under the control of endogenous regulatory elements and reducing risks associated with random insertion into the genome. Gene-specific targeting has historically been limited to mouse embryonic stem cells. The development of zinc finger nucleases (ZFNs) has permitted efficient genome editing in transformed and primary cells that were previously thought to be intractable to such genetic manipulation. In vitro, ZFNs have been shown to promote efficient genome editing via homology-directed repair by inducing a site-specific double-strand break (DSB) at a target locus, but it is unclear whether ZFNs can induce DSBs and stimulate genome editing at a clinically meaningful level in vivo. Here we show that ZFNs are able to induce DSBs efficiently when delivered directly to mouse liver and that, when co-delivered with an appropriately designed gene-targeting vector, they can stimulate gene replacement through both homology-directed and homology-independent targeted gene insertion at the ZFN-specified locus. The level of gene targeting achieved was sufficient to correct the prolonged clotting times in a mouse model of haemophilia B, and remained persistent after induced liver regeneration. Thus, ZFN-driven gene correction can be achieved in vivo, raising the possibility of genome editing as a viable strategy for the treatment of genetic disease.     

Disruption of mouse Slx4, a regulator of structure-specific nucleases, phenocopies Fanconi anemia

The evolutionarily conserved SLX4 protein, a key regulator of nucleases, is critical for DNA damage response. SLX4 nuclease complexes mediate repair during replication and can also resolve Holliday junctions formed during homologous recombination. Here we describe the phenotype of the Btbd12 knockout mouse, the mouse ortholog of SLX4, which recapitulates many key features of the human genetic illness Fanconi anemia. Btbd12-deficient animals are born at sub-Mendelian ratios, have greatly reduced fertility, are developmentally compromised and are prone to blood cytopenias. Btbd12(-/-) cells prematurely senesce, spontaneously accumulate damaged chromosomes and are particularly sensitive to DNA crosslinking agents. Genetic complementation reveals a crucial requirement for Btbd12 (also known as Slx4) to interact with the structure-specific endonuclease Xpf-Ercc1 to promote crosslink repair. The Btbd12 knockout mouse therefore establishes a disease model for Fanconi anemia and genetically links a regulator of nuclease incision complexes to the Fanconi anemia DNA crosslink repair pathway.     

Histone hydrolase in tadpole liver

Résumé Le cytoplasme du foie de têtard contient une enzyme qui hydrolyse spécifiquement les protéines basiques telle que la polylysine, la protamine et les histones. L'addition d'autres protéines telles que l'albumine, la globuline et l'acide polyglutamique inhibe l'activité de cette enzyme. Les résultats obtenus montrent que cette hydrolysase extraite du foie de têtard diffère des cathépsines ou de l'histone-hydrolase du rein de rat.

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This work was supported by research grants from the National Institute of Arthritis and Metabolic Diseases No. AM 09602, National Science Foundation No. GB-22663 and from the National Cancer Institute No. CA 07174.     

Highly mechanical and high-temperature properties of Cu–Cu joints using citrate-coated nanosized Ag paste in air

Today, a growing number of third-generation semiconductor-based power devices are used in products that can continuously operate at high temperatures for extended periods of time. Hence, traditional tin-lead and lead-free solders are no longer suitable for modern electronic packaging. A common method is to apply Ag paste for bare Cu–Cu joints under an inert or reductive atmosphere. In this study, the citrate-coated nanosized Ag paste was utilized to generate robust bare Cu–Cu joints under atmospheric conditions. The average size of citrate-coated Ag particles was approximately 4.76 ?nm after being cleaned by deionized water and acetone. The effects of process parameters, such as cleaning, joining temperature, holding time, and joining pressure, on the mechanical properties of the bare Cu–Cu joints were thoroughly investigated. Increasing washing and joining temperatures resulted in a shear strength increase of up to 28.2 ?MPa ?at a joining temperature of 260 ?°C after seven washes. In addition, a holding time of 30 ?min and a joining pressure of 1 ?MPa were selected as optimal process conditions for the application of citrate-coated nanosized Ag paste onto bare Cu–Cu joints. The newly developed Cu–Cu joints showed excellent thermal stability at 150 ?°C using the citrate-coated nanosized Ag paste. After long-term aging, the joints exhibited stability at 250 ?°C for 144 ?h, indicating a good high-temperature reliability for three-dimensional integrated circuits (3D ICs) fabricated under atmospheric conditions.     

Producer-consumer biomass in montane forests on the Arizona Mogollon Plateau

Normal 0 false false false EN-US X-NONE X-NONE MicrosoftInternetExplorer4 /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;} A substantially complete compilation of producer-consumer biomass was achieved for two montane forest reference stands on the Arizona Mogollon Plateau. This compilation, containing published and previously unpublished data, shows these ponderosa-pine-dominated stands to be near the lower end of the biomass range of commercial forest types. The two stands averaged approximately 75 metric tons/ha of plant biomass. Consumers made up less than 0.01 percent of the forest biomass. About 9/10 of the measured consumer biomass consisted of domestic and native ruminants.