Investigating Circular RNAs Using qRT-PCR; Roundup of Optimization and Processing Steps
Abstract
1. Introduction
2. Results
2.1. Database Annotation, Composition, and Sequence Characterization of circRNAs
2.2. Junction Sequence Mapping and Primer Design
2.3. Primer Validation
2.4. RNAse R Treatment Optimization and RNA Quality Control
2.5. Assessing Enrichment of circRNA following RNAse R Treatment
2.6. Database Prediction of Interacting miRNAs and Construction of circRNA-miRNA-mRNA Networks
3. Discussion
4. Materials and Methods
4.1. Cell Lines
4.2. RNA Extraction
4.3. RNAse R Treatment
4.4. RNA Clean-Up and Quantification
4.5. cDNA Synthesis
4.6. Primer Validation PCR and Agarose Gel Electrophoresis
4.7. qRT-PCR
4.8. Data Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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CircRNA | Parental Gene | CircBase ID | Structure | Genomic Location | Length |
---|---|---|---|---|---|
circFOXO3 | FOXO3 | hsa_circ_0006404 | Exon 2 | chr6:108984657-108986092 | 1435 bp |
circPVT1 | PVT1 lncRNA | hsa_circ_0009143 | Exon 2 | chr8:128867400-128903244 | 410 bp |
circANAPC2 | ANAPC2 | Unannotated variant | Exon 1 | chr9:137188416-137188560 | 145 bp |
circACAP2 | ACAP2 | Unannotated variant | Exon 3-1 | Chr3:195442795-195381903 | 404 bp |
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Drula, R.; Braicu, C.; Chira, S.; Berindan-Neagoe, I. Investigating Circular RNAs Using qRT-PCR; Roundup of Optimization and Processing Steps. Int. J. Mol. Sci. 2023, 24, 5721. https://doi.org/10.3390/ijms24065721
Drula R, Braicu C, Chira S, Berindan-Neagoe I. Investigating Circular RNAs Using qRT-PCR; Roundup of Optimization and Processing Steps. International Journal of Molecular Sciences. 2023; 24(6):5721. https://doi.org/10.3390/ijms24065721
Chicago/Turabian StyleDrula, Rares, Cornelia Braicu, Sergiu Chira, and Ioana Berindan-Neagoe. 2023. "Investigating Circular RNAs Using qRT-PCR; Roundup of Optimization and Processing Steps" International Journal of Molecular Sciences 24, no. 6: 5721. https://doi.org/10.3390/ijms24065721
APA StyleDrula, R., Braicu, C., Chira, S., & Berindan-Neagoe, I. (2023). Investigating Circular RNAs Using qRT-PCR; Roundup of Optimization and Processing Steps. International Journal of Molecular Sciences, 24(6), 5721. https://doi.org/10.3390/ijms24065721