how important a terminator is in cloning? - (Dec/03/2007 )
I want to move whole cassette from one vector to another, i chosed RE sites which are unique to both vectors. everything else seems good except the 89bp are missing from terminator sequence in cassette which i suppose to move.
should i do cloning with some bases of terminator missing and it will be safe or should i need to replace terminator, and if the second option works then how can i replace.
May be the terminator sequence is a modified one where 89 bases were removed purposely. What sort of terminator sequence do you have in the plasmid?
actually i wanna move whole cassette from agro-binary vector and will clone this in another binary vector. terminator is not missing but in fact the two enzymes im using are the only unique enzymes in this vector, and one of them lies inside the terminator sequence. someother site i cant use because it will make my cassette size horribly large......
Is PCR an option for you? I normally donot play around with regulatory elements in my vector unless absolutely necessary. If its just to clone a transgene, I might amplify it with desired sites and clone into vector of choice. Unless one is familiar with a vector, it is not advisable to meddle with regulatory elements.