Which is better? To blunt my insert and vector or use linkers? - To insert the 3.5 kb insert cut with EcoRI and SalI in to a (Feb/28/2005 )
To ligate the 3.5 kb DNA insert restricted with EcoRI and SalI in to a vector cut with EcoRI ,what would be better?
1. To blunt the ends of both insert and vector and ligate? or
2. To use linkers/adaptors to make SalI end of insert to EcoRI end ?
and if latter is better which is the best possible adaptor sequence? PLz help as this concern is bugging me a lot!!!! Thanx.
i suppose your fragment will look like Eco -------- Sal I
Hence, restriction sites are not compatible. That's why i would ligate this insert like a standard one.
but do you mean you want to change the sense of the restricion sites?
If you want to do so, you may blunt the ends and take an appropriate ligation protocol ( see topic 5292 at molecular biology) http://www.protocol-online.org/forums/inde...?showtopic=5292
hope i've helped you for a few...