Key result
Circular RNA expression vectors produce trans-spliced mRNAs that cause false-positive signals in IRES discovery screens.
Why the study?
Circular RNA expression vectors are used to identify and characterize RNA sequences with IRES activity, but potential confounding trans-spliced mRNA products have not been fully characterized.
Interpretations of IRES activity based solely on circRNA expression vectors should be made with caution due to the potential for false-positive signals from trans-spliced mRNAs.
Embedded promoters may generate trans-spliced artifacts in circRNA vectors; leaves open IRES assay specificity concerns.
Circular (circ) RNA expression vectors are used as a method of identifying and characterizing RNA sequences that harbor internal ribosome entry site (IRES) activity. During the course of developing a vector series tailored for IRES discovery, we found evidence for the occurrence of trans-spliced mRNAs arising when sequences with promoter activity were embedded between the upstream CTD and downstream NTD exons of the pre-mRNA. These trans-spliced products regenerate the same open reading frame expected from a circRNA and can lead to false-positive signals in screens relying on circRNA expression vectors for IRES discovery. Our results caution against interpretations of IRES activity solely based on results obtained from circRNA expression vectors.
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Chu et al. (2021) studied this question. circRNA expression vectors was evaluated on Occurrence of trans-spliced mRNAs. Circular RNA expression vectors can produce trans-spliced mRNAs that regenerate the expected open reading frame, leading to false-positive signals in internal ribosome entry site discovery screens.
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