Cre recombinase

Cre recombinase is a tyrosine recombinase enzyme derived from the P1 bacteriophage. It is widely used in genetic engineering to carry out site-specific recombination events. The enzyme recognizes specific DNA sequences known as loxP sites and catalyzes the recombination between these sites. This system is commonly referred to as the Cre-Lox recombination system.

Structure and Function

Cre recombinase is a 38 kDa protein that functions as a homotetramer. Each monomer of Cre recombinase binds to a 34 base pair loxP site, which consists of two 13 base pair palindromic sequences flanking an 8 base pair asymmetric core sequence. The enzyme mediates recombination by cleaving the DNA at the loxP sites and rejoining the DNA strands in a new configuration.

Mechanism of Action

The recombination process involves several steps: 1. **Binding**: Cre recombinase binds to the loxP sites. 2. **Cleavage**: The enzyme introduces a double-strand break at the loxP sites. 3. **Strand Exchange**: The DNA strands are exchanged between the loxP sites. 4. **Ligation**: The DNA strands are re-ligated, resulting in a recombined DNA sequence.

Applications

Cre recombinase is extensively used in molecular biology and genetics for various applications, including:

• **Conditional Gene Knockout**: By flanking a gene of interest with loxP sites, researchers can use Cre recombinase to delete the gene in specific tissues or at specific developmental stages.
• **Gene Activation**: Cre recombinase can be used to activate gene expression by removing a stop sequence flanked by loxP sites.
• **Chromosomal Rearrangements**: The enzyme can induce inversions, deletions, and translocations of chromosomal segments.

Cre-Lox System in Research

The Cre-Lox system is a powerful tool in mouse genetics and has been used to create various transgenic mouse models to study gene function and disease mechanisms. By using tissue-specific or inducible promoters to drive Cre expression, researchers can achieve precise control over gene recombination events.

Advantages and Limitations

Advantages

• **Specificity**: The Cre-Lox system is highly specific, allowing for targeted genetic modifications.
• **Versatility**: It can be used for a wide range of genetic manipulations, including gene knockouts, knock-ins, and chromosomal rearrangements.

Limitations

• **Efficiency**: The efficiency of recombination can vary depending on the accessibility of loxP sites and the expression levels of Cre recombinase.
• **Off-target Effects**: Although rare, off-target recombination events can occur, leading to unintended genetic modifications.

See Also

• Recombinase
• Genetic engineering
• Transgenic organism
• Gene knockout
• Mouse genetics