cloning help - (Feb/11/2011 )

Hi,

I would like to sub-clone apoA-5 cDNA cloned in pcDNA 3.1 into pAAV-MCS for that I am designing the primers for apoA-5 with BamHI and XbaI RE sites. It is suggested that for the good cloning insert should be in frame with the vector. What is the meaning of in frame. I will be grateful if anyone can explain me in detail with a good example.

vinu on Fri Feb 11 21:47:07 2011 said:

Vinu,

In frame means that the insert should be in the reading frame so that, when the insert is transcribed and then translated, it produces a meaningful protein, in your case, apoA-5.

gt_ameya on Sat Feb 12 05:15:45 2011 said:

Thanks for the information. I would also like to know how i can make that happen do i have to alter my primer sequence according to the vector to make it happen.

If your insert is a coding sequence (contains a start and a stop codon) , it should not matter. You only have to pay attention to the reading frame if you want to create a fusion protein for example.
Correct my if I am wrong