Questioning the utility of RNA-DNA chimera in gene repair.

Détails

ID Serval
serval:BIB_18377
Type
Article: article d'un périodique ou d'un magazine.
Sous-type
Lettre (letter): communication adressée à l'éditeur.
Collection
Publications
Institution
Titre
Questioning the utility of RNA-DNA chimera in gene repair.
Périodique
Science STKE
Auteur⸱e⸱s
Stasiak A.
ISSN
1945-0877
Statut éditorial
Publié
Date de publication
2001
Peer-reviewed
Oui
Volume
0ctober
Pages
N/A
Langue
anglais
Résumé
In principle, we should be glad that Eric Kmiec and his colleagues published in Science's STKE (1) a detailed experimental protocol of their gene repair method (2, 3). However, a careful reading of their contribution raises more doubts about the method. The research published in Science five years ago by Kmiec and his colleagues was said to demonstrate that chimeric RNA-DNA oligonucleotides could correct the mutation responsible for sickle cell anemia with 50% efficiency (4). Such a remarkable result prompted many laboratories to attempt to replicate the research or utilize the method on their own systems. However, if the method worked at all, which it rarely did, the achieved efficiency was usually lower by several orders of magnitude.
Now, in the Science's STKE protocol, we are given crucial information about the method and why it is so important to utilize these expensive chimeric RNA-DNA constructs. In the introduction we are told that the RNA-DNA duplex is more stable than a DNA-DNA duplex and so extends the half-life of the complexes formed between the targeted DNA and the chimeric RNA-DNA oligonucleotides. This logical explanation, however, conflicts with the statement in the section entitled "Transfection with Oligonucleotides and Plasmid DNA" that Kmiec and colleagues have recently demonstrated that classical single-stranded DNA oligonucleotides with a few protective phosphothioate linkages have a "gene repair conversion frequency rivaling that of the RNA/DNA chimera". Indeed, the research cited for that result actually states that single-stranded DNA oligonucleotides are in fact several-fold more efficient (3.7-fold) than the RNA-DNA chimeric constructs (5). If that is the case, it raises the question of why Kmiec and colleagues emphasize the importance of the RNA in their original chimeric constructs. Their own new results show that modified single-stranded DNA oligonucleotides are more effective than the expensive RNA-DNA hybrids. Moreover, the current efficiency of the gene repair by RNA-DNA hybrids, according to Kmiec and colleagues in their recent paper is only 4×10-4 even after several hours of pre-selection permitting multiplification of bacterial cells with the corrected plasmid (5). This efficiency is much lower than the 50% value reported five years ago, but is assuredly much closer to the reality.
Création de la notice
19/11/2007 12:12
Dernière modification de la notice
20/08/2019 12:48
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