Posted 24 July 2012 - 07:14 AM
I am cloning my gene of interest in pAAV-DIO vector to cell specific expression in brain. However, my gene doesn't present any kozak consensus sequence upstream and downstream of the ATG naturally. The Kozak consensus sequence plays a major role in the initiation of the translation process. Therefore, I can add kozak sequence upstream of ATG; however it is not possible in downstream. This manipulation might work. Even without the Kozak sequence, the translation efficiency of the protein might drop, or will drop. I do not have experience with the mammalian expression system. Is the Kozak sequence really needed for cloning?
Kindly suggest me.
Posted 24 July 2012 - 06:36 PM
Posted 25 July 2012 - 02:16 PM
I added GCCACC before ATG in the primer. I hope this should work.'