2 replies to this topic

[tomgar]

hi everyone
last time i asked for some tips about partial digestion, i forgot to give some details: i use 2 enzymes NcoI and HindIII, i cut pBluescript where my insert is put in. Generally I have 3 restriction sites in that insert: 2 for NcoI and 1 for HindIII.
What do You think about it?
How to manage that problem?
Thanks
Best
tomek

[mybbff]

Partial digestion is not a very good method to identify the recombinant clone. To this end, you may

1) reduce the digestion time;
2) reduce the restriction enzyme;

Also, you could choose other methods to complete the identification, such as sequencing, or restricting by other double=enzyme( one in the vector and another in the insert).

[tomgar]

mybbff, on Jan 19 2005, 08:33 PM, said:

Partial digestion is not a very good method to identify the recombinant clone. To this end, you may

1) reduce the digestion time;
2) reduce the restriction enzyme;

Also, you could choose other methods to complete the identification, such as sequencing, or restricting by other double=enzyme( one in the vector and another in the insert).

i have to make partial digestion to obtain some DNA fragment needed to forward subcloning, it is necessary step for me. I will check both methods, thanks.