I am wondering whether the length of the insert cloned in a lentivector would effect the packaging efficiency or the viral titer? Did any one notice any such changes?
Absolutely. There is a huge effect of the size of the insert vs titers.The longer the insert, the less virus you'll get. I have gotten virus to be produced from lentiviral vectors that are 12.5-13 kb in size (the size of the actual lenti, from LTR to LTR is then little over 10k). But titers are really low as compared to lentis with a vector size of ±9kb (lenti genome size of 6-7kb), several log that is. It does make sense, there is a limit of what size RNA you can fit into the virion...
To have efficient packaging, the region between the 5’and 3’LTR must not exceed 9kb, which is the size of the native HIV genome. To calculate the size of the insert that the lentivector can accommodate, download the vector sequence and determine the size of this region. The difference between 9kb and the size of the 5’- to 3’-LTR is the limit of the size of the insert for optimum packaging.
Hi Afzal,
The link between size of genome and titer is very well describe in this article:
http://online.liebertpub.com/doi/abs/10.1089/104303401753153947
If you don't have the access to this paper i can send you it by email if you want.
Nicolas
Hi Nicolas,
Could you please send me the article? My email address is [email protected]
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