What amount of insert:vector should I use if I want a 3:1 molar ratio? - (Apr/24/2006 )

Hi all,

I'm doing some cloning experiments now and I wanted to make sure how to calculate how much insert and vector I need. My insert is 110bp and my vector is 8kb and I want a 3:1 molar ratio of insert to vector. The way I was taught was that since my vector is 80 times bigger than my insert, I divide 3 by 80 (which is 0.0375) and that is how much more insert I need than vector. That's what I was told which I think is incorrect because that means the larger the vector is compared to the insert , the less insert we need. I thought it was the other way around? Can someone please confirm for me? So if I have 6ng/ul of insert and 10ng/ul of vector...how many ng of vector would I need per 6ng of insert? It's my first time, please help!

Selenilum

Here you have the formule given by Promega for pTarget. In this case, the size of the plasmid is 5.7kb (60ng/ul) and the insert 0.8kb. Make the appropriate changes for your experiments.




To do this, you need to know how much amount you got of each (vector and insert).

I hope this could help you.

Hi Gsamsa,

Thanks so much for your reply. I think that was the equation I used as well, just wanted to make sure.