Efficient delivery of Cre-recombinase to neurons in vivo and stable transduction  of neurons using adeno-associated and lentiviral vectors.

Ahmed, Bushra Y; Chakravarthy, Sridhara; Eggers, Ruben; Hermens, Wim T J M C; Zhang, Jing Ying; NICLOU, Simone P.; Levelt, Christiaan; Sablitzky, Fred; Anderson, Patrick N; Lieberman, A R; Verhaagen, Joost

doi:10.1186/1471-2202-5-4

Article (Scientific journals)

Efficient delivery of Cre-recombinase to neurons in vivo and stable transduction of neurons using adeno-associated and lentiviral vectors.

Ahmed, Bushra Y; Chakravarthy, Sridhara; Eggers, Ruben et al.

2004 • In BMC Neuroscience, 5, p. 4

Peer Reviewed verified by ORBi

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https://hdl.handle.net/10993/60260

DOI
10.1186/1471-2202-5-4

PubMed
15005815

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Keywords :

Luminescent Proteins; Recombinant Proteins; Viral Proteins; Green Fluorescent Proteins; EC 2.7.7.- (Cre recombinase); EC 2.7.7.- (Integrases); Animals; Cell Line; Dependovirus/genetics; Gene Expression; Gene Transfer Techniques; Genes, Reporter; Genetic Vectors/administration & dosage/genetics; Hippocampus/metabolism/virology; Integrases/administration & dosage/genetics/metabolism; Lentivirus/genetics; Luminescent Proteins/biosynthesis/genetics; Mice; Mice, Transgenic; Neocortex/metabolism/virology; Neurons/metabolism/virology; Recombinant Proteins/genetics/metabolism; Recombination, Genetic/drug effects/genetics; Transgenes; Viral Proteins/administration & dosage/genetics/metabolism

Abstract :

[en] BACKGROUND: Inactivating genes in vivo is an important technique for establishing their function in the adult nervous system. Unfortunately, conventional knockout mice may suffer from several limitations including embryonic or perinatal lethality and the compensatory regulation of other genes. One approach to producing conditional activation or inactivation of genes involves the use of Cre recombinase to remove loxP-flanked segments of DNA. We have studied the effects of delivering Cre to the hippocampus and neocortex of adult mice by injecting replication-deficient adeno-associated virus (AAV) and lentiviral (LV) vectors into discrete regions of the forebrain. RESULTS: Recombinant AAV-Cre, AAV-GFP (green fluorescent protein) and LV-Cre-EGFP (enhanced GFP) were made with the transgene controlled by the cytomegalovirus promoter. Infecting 293T cells in vitro with AAV-Cre and LV-Cre-EGFP resulted in transduction of most cells as shown by GFP fluorescence and Cre immunoreactivity. Injections of submicrolitre quantities of LV-Cre-EGFP and mixtures of AAV-Cre with AAV-GFP into the neocortex and hippocampus of adult Rosa26 reporter mice resulted in strong Cre and GFP expression in the dentate gyrus and moderate to strong labelling in specific regions of the hippocampus and in the neocortex, mainly in neurons. The pattern of expression of Cre and GFP obtained with AAV and LV vectors was very similar. X-gal staining showed that Cre-mediated recombination had occurred in neurons in the same regions of the brain, starting at 3 days post-injection. No obvious toxic effects of Cre expression were detected even after four weeks post-injection. CONCLUSION: AAV and LV vectors are capable of delivering Cre to neurons in discrete regions of the adult mouse brain and producing recombination.

Disciplines :

Oncology

Author, co-author :

Ahmed, Bushra Y; Department of Anatomy and Developmental Biology, University College London, Gower Street, London WC1E 6BT, UK. Bushra.Ahmed@ucl.ac.uk

Chakravarthy, Sridhara

Eggers, Ruben

Hermens, Wim T J M C

Zhang, Jing Ying

NICLOU, Simone P. ; Netherlands Institute for Brain Research, Amsterdam, The Netherlands

Levelt, Christiaan

Sablitzky, Fred

Anderson, Patrick N

Lieberman, A R

Verhaagen, Joost

External co-authors :

yes

Language :

English

Title :

Efficient delivery of Cre-recombinase to neurons in vivo and stable transduction of neurons using adeno-associated and lentiviral vectors.

Publication date :

30 January 2004

Journal title :

BMC Neuroscience

eISSN :

1471-2202

Publisher :

BioMed Central, United Kingdom

Volume :

Pages :

Peer reviewed :

Peer Reviewed verified by ORBi

Available on ORBilu :

since 19 February 2024

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