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Engineering a ribozyme cleavage-induced split fluorescent aptamer complementation assay
- Source :
- Nucleic Acids Research, 44 (10), Nucleic Acids Research
- Publication Year :
- 2016
- Publisher :
- Oxford University Press, 2016.
-
Abstract
- Hammerhead ribozymes are self-cleaving RNA molecules capable of regulating gene expression in living cells. Their cleavage performance is strongly influenced by intra-molecular loop–loop interactions, a feature not readily accessible through modern prediction algorithms. Ribozyme engineering and efficient implementation of ribozyme-based genetic switches requires detailed knowledge of individual self-cleavage performances. By rational design, we devised fluorescent aptamer-ribozyme RNA architectures that allow for the real-time measurement of ribozyme self-cleavage activity in vitro. The engineered nucleic acid molecules implement a split Spinach aptamer sequence that is made accessible for strand displacement upon ribozyme self-cleavage, thereby complementing the fluorescent Spinach aptamer. This fully RNA-based ribozyme performance assay correlates ribozyme cleavage activity with Spinach fluorescence to provide a rapid and straightforward technology for the validation of loop–loop interactions in hammerhead ribozymes.<br />Nucleic Acids Research, 44 (10)<br />ISSN:1362-4962<br />ISSN:0301-5610
- Subjects :
- 0301 basic medicine
biology
010405 organic chemistry
Aptamer
Genetic Complementation Test
Ribozyme
RNA
Aptamers, Nucleotide
01 natural sciences
0104 chemical sciences
03 medical and health sciences
030104 developmental biology
Biochemistry
Protein-fragment complementation assay
Genetics
biology.protein
Methods Online
RNA, Catalytic
Mammalian CPEB3 ribozyme
Genetic Engineering
Hairpin ribozyme
VS ribozyme
Ligase ribozyme
Fluorescent Dyes
Subjects
Details
- Language :
- English
- ISSN :
- 13624962 and 03015610
- Database :
- OpenAIRE
- Journal :
- Nucleic Acids Research, 44 (10), Nucleic Acids Research
- Accession number :
- edsair.doi.dedup.....5b8d48a0cf756e08cd3f7c33bf7bbcc3