Optimizing approaches for targeted integration of transgenic cassettes by integrase-mediated cassette exchange in mouse and human stem cells.

To enable robust expression of transgenes in stem cells, recombinase-mediated cassette exchange at safe harbor loci is frequently adopted. The choice of recombinase enzyme is a critical parameter to ensure maximum efficiency and accuracy of the integration event. We have explored the serine recombinase family of site-specific integrases and have directly compared the efficiency of PhiC31, W-beta, and Bxb1 integrase for targeted transgene integration at the Gt(ROSA)26Sor locus in mouse embryonic stem cells.

Rangers on the frontline of wildlife monitoring: a case study on African lions in Uganda's Nile Delta.

Regular population monitoring of imperilled charismatic species such as large carnivores is critical for conservation. However, the role of monitoring in conservation is frequently diminished due to: 1) surveys being implemented in isolation, 2) limited on-ground-capacity leading to infrequent monitoring, and 3) inappropriate methods being applied. Wildlife monitoring is often resource-intensive and the utility and cost of different field protocols is rarely reported.

Shieldin and CST co-orchestrate DNA polymerase-dependent tailed-end joining reactions independently of 53BP1-governed repair pathway choice.

Tumor suppressor p53-binding protein 1 (53BP1) regulates DNA end joining in lymphocytes, diversifying immune antigen receptors. This involves nucleosome-bound 53BP1 at DNA double-stranded breaks (DSBs) recruiting Rap1-interacting factor 1 homolog (RIF1) and shieldin, a poorly understood DNA-binding complex. The 53BP1-RIF1-shieldin axis is pathological in BRCA1-mutated cancers, blocking homologous recombination (HR) and driving illegitimate nonhomologous end joining (NHEJ).

Mutagenesis on a complex mouse genetic background by site-specific nucleases.

Mouse models with complex genetic backgrounds are increasingly used in preclinical research to accurately model human disease and to enable temporal and cell-specific evaluation of genetic manipulations. Backcrossing mice onto these complex genetic backgrounds takes time and leads to significant wastage of animals. In this study, we aimed to evaluate whether site-specific nucleases could be used to generate additional genetic mutations in a complex genetic background, using the REVERSA mouse model of atherosclerosis, a model harbouring four genetically altered alleles.