Bibliographic Details
| Title: |
RETRACTED ARTICLE: RNA editing of BFP, a point mutant of GFP, using artificial APOBEC1 deaminase to restore the genetic code. |
| Authors: |
Bhakta, Sonali1 (AUTHOR), Sakari, Matomo1 (AUTHOR), Tsukahara, Toshifumi1 (AUTHOR) tukahara@jaist.ac.jp |
| Source: |
Scientific Reports. 2/14/2025, Vol. 15 Issue 1, p1-13. 13p. |
| Subject Terms: |
*HAIRPIN (Genetics), *LIFE sciences, *CYTOLOGY, *RNA editing, *SYNTHETIC enzymes |
| Abstract: |
Many genetic diseases are caused by T-to-C point mutations. Hence, editing of mutated genes represents a promising strategy for treating these disorders. We engineered an artificial RNA editase by combining the deaminase domain of APOBEC1 (apolipoprotein B mRNA editing catalytic polypeptide 1) with a guideRNA (gRNA) which is complementary to target mRNA. In this artificial enzyme system, gRNA is bound to MS2 stem-loop, and deaminase domain, which has the ability to convert mutated target nucleotide C-to-U, is fused to MS2 coat protein. As a target RNA, we used RNA encoding blue fluorescent protein (BFP) which was derived from the gene encoding GFP by 199 T > C mutation. Upon transient expression of both components (deaminase and gRNA), we observed GFP by confocal microscopy, indicating that mutated 199C in BFP had been converted to U, restoring original sequence of GFP. This result was confirmed by PCR–RFLP and Sanger's sequencing using cDNA from transfected cells, revealing an editing efficiency of approximately 21%. Although deep RNA sequencing result showed some off-target editing events in this system, we successfully developed an artificial RNA editing system using artificial deaminase (APOBEC1) in combination with MS2 system could lead to therapies that treat genetic disease by restoring wild-type sequence at the mRNA level. [ABSTRACT FROM AUTHOR] |
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