Single promoter system for making sirna expression cassettes...

C - Chemistry – Metallurgy – 12 – N

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C12N 15/63 (2006.01) C12N 15/10 (2006.01) C12N 15/11 (2006.01) C12N 15/66 (2006.01)

Patent

CA 2493251

The present invention relates to methods and compositions for the elucidation of gene function and the identification of novel genes. Specifically, the present invention relates to methods and compositions for improved functional genomic screening, functional inactivation of specific essential or non- essential genes, and identification of genes that are modulated in response to specific stimuli or encode recognizable phenotypic traits. In particular, the compositions of the present invention include, but are not limited to, expression cassettes comprising a novel polymerase primer hairpin linker that allows rapid construction of a single transcriptional unit encoding both strands of a hairpin siRNA, regardless of sequence. In addition, the present invention includes libraries comprising the expression cassettes of the invention, including vectors for transforming cells, such as replication- deficient retroviral vectors. The invention also includes methods for the production and screening of siRNA libraries, as well as therapeutic uses for the siRNAs expressed in accordance with the invention.

Méthodes et compositions permettant d'élucider la fonction génique et d'identifier de nouveaux gènes. Spécifiquement, la présente invention concerne des méthodes et des compositions permettant d'améliorer le criblage génomique fonctionnel, l'inactivation fonctionnelle de gènes essentiels ou non essentiels spécifiques et l'identification de gènes qui sont modulés en réponse à des stimuli spécifiques ou qui codent des traits phénotypiques reconnaissables. En particulier, les compositions selon la présente invention comportent, entre autres, des cassettes d'expression contenant un nouveau lieur en épingle à cheveux d'amorce de polymérase qui permet une construction rapide d'une unité transcriptionnelle unique codant les deux brins d'ARNsi en épingle à cheveux, indépendamment de la séquence. De plus, la présente invention concerne des génothèques contenant des cassettes d'expression selon la présente invention, dont des vecteurs de transformation de cellules, tels que des vecteurs rétroviraux à déficience de réplication. La présente invention concerne encore des méthodes de production et de criblage de génothèques d'ARNsi, ainsi que les utilisations thérapeutiques des ARNsi exprimés selon la présente invention.

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