Re: Ligation Problem# Biology - 生物学
g*m
1 楼
man, refer to gzip's post.
the ligation is somehow vulnerable.
1. check with your lab fellow to make sure your ligase is still alive.
2. always use latest ligation buffer, you can even smell the thio stuff if you
are using NEB enzyme.
3. critical thing is your insert: never directly use the insert purified from
gel, you need gel purification followed by ethanol precipitation to make sure
the purity. However, you can use gel extracted vector for ligation in small
volume.
4. add enough insert, for
the ligation is somehow vulnerable.
1. check with your lab fellow to make sure your ligase is still alive.
2. always use latest ligation buffer, you can even smell the thio stuff if you
are using NEB enzyme.
3. critical thing is your insert: never directly use the insert purified from
gel, you need gel purification followed by ethanol precipitation to make sure
the purity. However, you can use gel extracted vector for ligation in small
volume.
4. add enough insert, for