Preparation of sequence libraries from non-denatured rna and...

C - Chemistry – Metallurgy – 12 – N

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C12N 15/10 (2006.01) C12N 15/11 (2006.01) C12Q 1/68 (2006.01)

Patent

CA 2392959

A method is provided for preparing libraries of DNA sequences from non- denatured RNA. In one embodiment, the method includes: forming a library of target RNA fragments by contacting multiple copies of non-denatured target RNA sequences with a library of random oligonucleotides in the presence of a hydrolytic agent under conditions such that a subgroup of the library of random oligonucleotides hybridizes to the target RNA, whereupon the hydrolytic agent hydrolyzes the target RNA at a site near the 5' end of each hybridized random oligonucleotide and wherein the 3' ends of each fragment contains the entire sequence to which a random oligonucleotide in the subgroup hybridized; and forming a library of templates for primer extension.

L'invention concerne un procédé de préparation de bibliothèques de séquences d'ADN à partir d'ARN non-dénaturé. Dans un mode de réalisation, ce procédé consiste à former une bibliothèque de fragments d'ARN cible par mise en contact de plusieurs copies de séquences d'ARN cible non-dénaturé avec une bibliothèque d'oligonucléotides aléatoires en présence d'un agent hydrolytique sous des conditions telles qu'un sous-groupe de la bibliothèque d'oligonucléotides aléatoires s'hybride sur l'ARN cible, l'agent hydrolytique hydrolysant l'ARN cible sur un site proche de l'extrémité 5' de chaque oligonucléotide aléatoire hybridé, et chaque extrémité 3' de chaque fragment contenant la séquence entière sur laquelle un oligonucléotide aléatoire du sous-groupe s'est hybridée. Ce procédé consiste également à former une bibliothèque de modèles pour extension de l'amorce.

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