topo cloning for sequencing? - (Sep/12/2005 )
ciao!I'm trying sequencing some bisulphite modified DNA and I read that it is better to sequence using the topo-cloning system....so I'm searching for a good but CHEAP topo cloning system..you usually utilize topo cloning and wich topo-cloning do you utilize? thank you all!
I was wondering where you read this information from?
TA cloning works just as well provided you are using converntional Taq that leaves overhangs. I am sure such a system is much cheaper than a TOPO kit.
As far as I know, the cheapest method is the one that uses home-made T-vector (Marchuk, D., et al., 1991, Nucl. Acids Res. 19(5), pp 1154).
I really recommend buying the legit vector from invitrogen. It you want to save money, you can dilte the topo vector about 5X (just use the proper buffer) and it will still work well. This can get you more reactions per purchase. Also, it you buy the ultra-competent cells, split the cells into multiple tubes for transformation and you will get more bang for your buck. This has worked well work me.