Methods comprising apoptosis inhibitors for the generation of transgenic pigs

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Stem Cell Related Patent Number US5624824

Title:Targeted cleavage of RNA using eukaryotic ribonuclease P and external guide sequence
Inventors:Yuan, Yan; New Haven, CT, USA
Guerrier-Takada, Cecilia; New Haven, CT, USA
Altman, Sidney; Hamden, CT, USA
Liu, Fenyong; New Haven, CT, USA
Summary:Described herein is a composition for targeting an RNA substrate for cleavage by eukaryotic RNAase P via an oligonucleotide. The invention relates to a target recognition sequence and a RNAase P binding sequence in the targeting of RNA by RNAase P such that a suitable oligoribonucleotide (designated “external guide sequence”, or EGS) is employed to form a hybrid with the target RNA, thereby creating a substrate for cleavage by RNAase P in vitro, after which the EGS hydrogen bonds to the targeted RNA to form a partial tRNA-like structure including the aminoacyl acceptor stem, the T stem and loop, and part of the D stem. Claims of the invention include the EGS in which the anticodon stem and loop was deleted as the most efficient EGS with human RNAase P. Further disclosed are methods by which modifications may be made within the T-loop, and methods by which a suitable EGS may be randomly selected and expressed in vivo to make a selected RNA a target for cleavage by the host cell RNAase P, thus preventing expression of the function of the target RNA. Therapeutic applications are included for the treatment of cancer as well as of viral and bacterial infections.
Abstract:It has been discovered that any RNA can be targeted for cleavage by RNAase P from eukaryotic cells, for example, human RNAase P, using a suitably designed oligoribonucleotide (external guide sequence, or EGS) to form a hybrid with the target RNA, thereby creating a substrate for cleavage by RNAase P in vitro. The EGS hydrogen bonds to the targeted RNA to form a partial tRNA like structure including the aminoacyl acceptor stem, the T stem and loop, and part of the D stem. The most efficient EGS with human RNAase P is the EGS in which the anticodon stem and loop was deleted. Modifications can also be made within the T-loop. Methods are also disclosed to randomly select and to express a suitable EGS in vivo to make a selected RNA a target for cleavage by the host cell RNAase P, thus preventing expression of the function of the target RNA. The methods and compositions should be useful to prevent the expression of disease-causing genes in vivo.
US Patent Website:Click Here for Full Text of Patent
Title Number:US5624824
Application Number:US1994000207547
Date Filed:07/03/1994
Date Published:29/04/1997
Assignee:Yale University, New Haven, CT, USA


 
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