Cre-mediated site-specific recombination has become an invaluable tool for manipulation of the murine genome. The ability to conditionally activate gene expression or to generate chromosomal alterations with this same tool would greatly enhance zebrafish genetics. This study demonstrates that the HSP70 promoter can be used to inducibly control expression of an enhanced green fluorescent protein (EGFP) -Cre fusion protein. The EGFP-Cre fusion protein is capable of promoting recombination between lox sites in injected plasmids or in stably inherited transgenes as early as 2 hr post-heat shock induction. Finally, the levels of Cre expression achieved in a transgenic fish line carrying the HSP70-EGFP-cre transgene are compatible with viability and both male and female transgenic fish are fertile subsequent to induction of EGFP-Cre expression. Hence, our data suggests that Cre-mediated recombination is a viable means of manipulating gene expression in zebrafish.
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http://dx.doi.org/10.1002/dvdy.20475 | DOI Listing |
Genome Biol
May 2024
College of Agronomy, Anhui Agricultural University, Hefei, 230036, People's Republic of China.
Background: High-efficiency prime editing (PE) is desirable for precise genome manipulation. The activity of mammalian PE systems can be largely improved by inhibiting DNA mismatch repair by coexpressing a dominant-negative variant of MLH1. However, this strategy has not been widely used for PE optimization in plants, possibly because of its less conspicuous effects and inconsistent performance at different sites.
View Article and Find Full Text PDFbioRxiv
February 2024
Department of Genome Sciences, University of Washington, Seattle, WA, USA.
The functional consequences of structural variants (SVs) in mammalian genomes are challenging to study. This is due to several factors, including: 1) their numerical paucity relative to other forms of standing genetic variation such as single nucleotide variants (SNVs) and short insertions or deletions (indels); 2) the fact that a single SV can involve and potentially impact the function of more than one gene and/or regulatory element; and 3) the relative immaturity of methods to generate and map SVs, either randomly or in targeted fashion, in or model systems. Towards addressing these challenges, we developed , a straightforward method that enables the multiplex generation and mapping of several major forms of SVs (deletions, inversions, translocations) throughout a mammalian genome.
View Article and Find Full Text PDFFront Neurosci
August 2023
Molecular Neurogenetics, Max Planck Institute of Psychiatry, Munich, Germany.
Corticotropin-releasing hormone expressing (CRH) neurons are distributed throughout the brain and play a crucial role in shaping the stress responses. Mouse models expressing site-specific recombinases (SSRs) or reporter genes are important tools providing genetic access to defined cell types and have been widely used to address CRH neurons and connected brain circuits. Here, we investigated a recently generated driver line expanding the CRH system-related tool box.
View Article and Find Full Text PDFJ Microbiol Methods
September 2022
Ajinomoto-Genetika Research Institute, 117545 Moscow, Russian Federation; Biological Department, Lomonosov Moscow State University, 119991 Moscow, Russian Federation.
A novel genome editing method for repeated introduction of foreign DNA, including insertion of rather large DNA fragments, into predesigned points in the Corynebacterium glutamicum chromosome was developed. The method is based on the implementation of the Dual-In/Out strategy, which was previously provided in Escherichia coli according to recombineering-based methods (Minaeva et al., 2008) and allowed step-by-step construction of marker-less plasmid free recombinant strains.
View Article and Find Full Text PDFTheor Appl Genet
July 2022
Plant Gene Engineering Center, Chinese Academy of Sciences Key Laboratory of South China Agricultural Plant Molecular Analysis and Genetic Improvement, Guangdong Key Laboratory of Applied Botany, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, 510650, China.
N-cre and C-cre added in separate lines reassemble functional Cre in F1 progeny to excise unnecessary DNA, including cre DNA, thereby eliminating generations needed to cross in and out cre. Crop improvement via transgenesis can benefit through efficient DNA integration strategies. As new traits are developed, new transgenes can be stacked by in planta site-specific integration near previous transgenes, thereby facilitating their introgression to field cultivars as a single segregation locus.
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