Sequencing of Plasmids - (Feb/01/2005 )

Hi! How do you sequence an insert in a Plasmid? Do you make a nick? How? Via Enzymes or in an ultrasoundbath?
How much ng of the plasmid do you use for the squencing reaction?

You can use the supercoiled plasmid, you don't need to cut it. As few as 10-50 ng are enough, but I usually add 1 ul straight of the miniprep, without even OD'ed it .

badcell is right. no need to do anything with the plasmid. Sequencing is like a PCR reaction only with just one primer.