I want to synthesise mRNA using in vitro transcription. I am trying to the best way to design my cDNA template to be used for this purpose. I read lot of papers where some 5' and 3' UTR sequences are added to the coding sequence to enhance the stability of the mRNA. For example these sequences from human beta globin genes are often used. Can someone tell me howto chose these sequences ?
Hello Dolon.
Go to NCBI, type your gene name and complete mRNA. It will give a full transcript sequence. Clone the full sequence of the gene containing ~70bp 5'-UTR and possible length of 3'-UTR by PCR using specific primer pairs containing the sequences with your full cDNA generated from total RNA.
Isolate and that will be used for your full transcript seq. containing 5'- and 3'-UTRs.
Good luck.
Will using beta globin gene UTRs instead of my genes specific UTR enhance my protein expression. My aim is to maximize mRNA stability and maximize protein expression?
The short answer is Yes. However, if you know that the UTRs in your mRNA of interest decrease the stability, it may be helpful.
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Biology
- Biomolecules
- Nucleic Acids
- DNA