Part:BBa_K4636033:Design
RCA_45_1802_2
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
After conducting literature reviews, we found that the majority of researchers use random primers, and no guidelines were found for designing RCA primers. Our secondary PI mentioned that designing primers independently can enhance specificity. Besides, if the Tm value of primer is close to our enzyme’s reaction temperature, we don’t need to anneal our primer with sample at lower temperature first.
Primer design primarily involves using SnapGene software[2] to manually search for primers on the complementary insert sequence (BBa_K46360040 & BBa_K46360041) that have Tm values close to the enzyme’s reaction temperature and GC content around 50%. These primers are then individually tested using the IDT OligoAnalyzer™ Tool[3], with any primers prone to self-dimer formation being discarded. Finally, the remaining primer candidates are tested against each other using the IDT OligoAnalyzer™ Tool to assess the possibility of forming hetero primer.
Source
No
References
1. Boss, M., & Arenz, C. (2020). A Fast and Easy Method for Specific Detection of Circular RNA by Rolling-Circle Amplification. *Chembiochem : a European journal of chemical biology*, *21*(6), 793–796. https://doi.org/10.1002/cbic.201900514
2. https://www.snapgene.com/
3. https://sg.idtdna.com/pages/tools/oligoanalyzer