Recombinase AAV Particles in Animal Experiments and Neurological Applications

The successful execution of neurobiology studies requires methods that are highly targetable, efficient, and precise. The arsenal of new sensors, actuators, recombinases, genes, RNA and base editing enzymes, and other genetically encoded tools for studying the nervous system is rapidly growing. Adeno-associated virus (AAV) vectors remain the most versatile and powerful approach for delivering these tools to the central nervous system (CNS). Among them, AAV is usually combined with Cre/loxP and Flp recombinase technologies to deliver loxP-dependent recombinases encoded by the Cre gene and has been widely used in the preparation of transgenic mice and various screening assays.

Cre Recombinase and LoxP Sites

Cre Recombination Enzyme (Cre) catalyzes recombination between LoxP sites, which consists of two 13bp reverse palindromic sequences and an 8bp intervening spacer sequence, the reverse palindromic sequence is the recognition and binding region of Cre Recombination Enzyme, and the spacer sequence determines the orientation of the LoxP sequence. When two LoxP loci are present in the cell genome, the Cre enzyme performs knockdown, insertion, flipping and translocation of the specific loci according to the orientation and position of the two loxP loci.

Cre/LoxP Recombination System induces Genetic Recombination

Several ways of inducing recombination exist in the Cre/LoxP system, which is based on the interaction of Cre recombinase with the loxP site.

(1) Two loxP sites are located on the same DNA strand and in the same orientation, and Cre recombinase knocks out the sequence between the loxPs;

(2) Two loxP sites are located on the same DNA strand and in opposite directions, Cre recombinase induces sequence flip between loxPs;

(3) Two loxP sites are located on different DNA strands or chromosomes, and Cre recombinase induces an exchange between the two DNA strands or chromosomal translocation;

(4) Four loxP sites are located on two DNA strands or chromosomes, and Cre recombinase induces sequence interchange between loxPs.

Recombinase AAV Particles

With the advantages of the Cre-LoxP recombinant system's specificity and high recombination efficiency, Creative Biogene scientists have developed a series of recombinase AAV particles by combining the advantages of the system with the serotype and promoter specificity of the AAV viral vector. PCR to confirm transfer plasmids and purity assessment. We offer recombinase AAV particles that can be flexibly and efficiently used to construct conditional knockout/overexpression mouse models, avoiding the mouse hybridization aspect of traditional transgenic animal construction, and can easily and efficiently support neurological related studies.

Some of our products are listed below, but please feel free to contact us for more information on any recombinase AAV particles you may be interested in.

Recombinase AAV Particles
Cre-GFP Adeno-associated virus(AAV Serotype 9)	Cre-GFP Adeno-associated virus(AAV Serotype 8)
Cre-GFP Adeno-associated virus(AAV Serotype 6)	Cre-GFP Adeno-associated virus(AAV Serotype 5)
Cre-GFP Adeno-associated virus(AAV Serotype 2)	Cre-GFP Adeno-associated virus(AAV Serotype 1)
Cre Adeno-associated virus(AAV Serotype 9)	Cre Adeno-associated virus(AAV Serotype 8)
Cre Adeno-associated virus(AAV Serotype 6)	Cre Adeno-associated virus(AAV Serotype 2)
Cre Adeno-associated virus(AAV Serotype 1)	Cre Adeno-associated virus(AAV Serotype 5)

Application of Cre/LoxP recombinant system in the construction of transgenic animals

Adeno-associated viruses (AAVs) are a commonly used tool in neuroscience to efficiently label, trace, and/or manipulate neuronal populations. Highly specific targeting can be achieved through recombinase-dependent AAVs in combination with transgenic rodent lines that express Cre-recombinase in specific cell types. Visualization of viral expression is typically achieved through fluorescent reporter proteins (e.g., GFP or mCherry) packaged within the AAV genome. For example, researchers injected Cre-dependent (AAV Serotype 5, AAV Serotype 8) into wild-type C57BL/6J mice and found Cre-independent viral expression of AAV encoding mCherry, GFP, or hM3Dq after immunohistochemical amplification of fluorescent reporter proteins.

Fig. 1 Antibody amplification of Cre-dependent viral expression. (Botterill, et al., 2021)

Thus, using the Cre/loxP system, discrete cell populations can be targeted by a combination of transgenic mice and viral injections. Using this approach, rodents are genetically modified to express Cre in specific cell types, which can be widely used in neuroscience research.

References:

Botterill, J.J.; et al. Off-Target Expression of Cre-Dependent Adeno-Associated Viruses in Wild-Type C57BL/6J Mice. eNeuro. 2021 Nov 24;8(6): ENEURO. 0363-21.2021.
Haery, L.; et al. Adeno-Associated Virus Technologies and Methods for Targeted Neuronal Manipulation. Front Neuroanat. 2019 Nov 26; 13:93.
Szeberényi, J. Problem-solving test: conditional gene targeting using the Cre/loxP recombination system. Biochem Mol Biol Educ. 2013 Nov-Dec;41(6):445-9.
Grames, M.S.; et al. Cre-dependent AAV vectors for highly targeted expression of disease-related proteins and neurodegeneration in the substantia nigra. FASEB J. 2018 Aug;32(8):4420-4427.

* For research use only. Not intended for any clinical use.

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Recombinase AAV Particles in Animal Experiments and Neurological Applications

Cre Recombinase and LoxP Sites

Cre/LoxP Recombination System induces Genetic Recombination

Recombinase AAV Particles

Application of Cre/LoxP recombinant system in the construction of transgenic animals

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