We describe a transgenesis platform for Drosophila melanogaster that integrates three recently developed technologies: a conditionally amplifiable bacterial artificial chromosome (BAC), recombineering, and bacteriophage PhiC31-mediated transgenesis. The BAC is maintained at low copy number, facilitating plasmid maintenance and recombineering, but is induced to high copy number for plasmid isola...

UNLABELLED Recombineering, in vivo genetic engineering with bacteriophage homologous recombination systems, is a powerful technique for making genetic modifications in bacteria. Two systems widely used in Escherichia coli are the Red system from phage λ and RecET from the defective Rac prophage. We investigated the in vivo dependence of recombineering on DNA replication of the recombining subst...

Temperature effects on spectral properties of the two types of rod photoreceptors in toad retina, "red" and "green" rods, were studied in the range 0-38 degrees C. Absorbance spectra of the visual pigments were recorded by single-cell microspectrophotometry (MSP) and spectral sensitivities of red rods were measured by electroretinogram (ERG) recording across the isolated retina. The red-rod vis...

OBJECTIVE To evaluate, using histological analysis, the systemic action and repair process of wounds produced on the back of rats and treated with red, infrared, or both lasers applied directly or indirectly to the wounds. BACKGROUND DATA Skin tissue repair and wound healing are complex processes that involve a series of dynamic events. Many benefits are associated with biomodulation using la...

The distribution of crossovers along unreplicated chromosomes of bacteriophage lambda has been examined by determining the density distributions and genotypes of particles in the progenies of crosses of density-labeled by ordinary parents in the presence of genetic blocks to replication. The Red and Rec systems combined produce crossovers primarily near the ends (especially the right end) of th...

Functional analysis of mammalian genes in vivo is primarily achieved through analysing knockout mice. Now that the sequencing of several mammalian genomes has been completed, understanding functions of all the genes represents the next major challenge in the post-genome era. Generation of knockout mutant mice has currently been achieved by many research groups but only by making individual knoc...

Gene targeting refers to the precise modification of a genetic locus using homologous recombination. The generation of novel cell lines and transgenic mouse models using this method necessitates the construction of a 'targeting' vector, which contains homologous DNA sequences to the target gene, and has for many years been a limiting step in the process. Vector construction can be performed in ...

Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas12a (Cpf1) has emerged as an effective genome editing tool in many organisms. Here, we developed and optimized a CRISPR-Cas12a-assisted recombineering system to facilitate genetic manipulation in bacteria. Using this system, point mutations, deletions, insertions, and gene replacements can be easily generated on the chromosome...

Recombineering is a widely-used approach to delete genes, introduce insertions and point mutations, and introduce epitope tags into bacterial chromosomes. Many recombineering methods have been described, for a wide range of bacterial species. These methods are often limited by (i) low efficiency, and/or (ii) introduction of "scar" DNA into the chromosome. Here, we describe a rapid, efficient, P...

To examine bacteriophage recombination in vivo, independent of such other processes as replication and packaging, substituted lambda phages bearing restriction site polymorphisms were employed in a direct physical assay. Bacteria were infected with two phage variants; DNA was extracted from the infected cells and cut with a restriction endonuclease. The production of a unique recombinant fragme...