if you are using a restriction enzyme of unknown type1 look on the tube it was sold in. They are always labelled. Alternatively ask your PI

If you have an unlabelled tube either use another enzyme that cuts you plasmid or test cut lambda dna or similar and see what sizes occur. This should give you a rough idea which enzyme it is then you can pcr a specific sequence of dna and cut it to prove which enzyme it is.

The probable reason that sequencing did not clarify this is that not everything was sequenced because the primers were inside of the cut site .or only designed to sequence a region of interest which did not include the cut site

Hi.....If you cut your plasmid with your unknown type enzyme and one of the unique restriction enzymes in your plasmid, based on restriction pattern i.e. size of release will guide you to find a clevage site of your unknown enzyme followed by sequencing and  if you have clear plasmid map.  You can also predict recognition seq of enzyme  by going with definitions of types of restriction enzymes. Further you can design test prove that based on deletion based mutant of plasmid.

Thanks for the replies, Paul and Sachin. My enzyme is not a restriction enzyme and I believe it could be some kind of nuclease. If primers were inside the cut site one will expect the sequencing reaction to fail and in my case i got the sequence full pUC19. five primers were designed cover pUC19. Sachin, I did your experiment with four unique enzymes spread across pUC19 and resulting pattern was same. I couldn't see any released product. So I couldn't  go further. 

I may be misunderstanding what you are doing but if you cut with most nucleases then the cutting is random and a huge variety of different lengths of linearised plasmid will be produced. For any given sequencing primer there will be a large number of fragments it will anneal to ....some longer than others but the sequencing will not distinguish between  all of these fragments so a longer fragment will sequence past the termination point of a shorter fragment and the sequence will just look like that of the longer fragment ( possibly with the peak intensities of the shorter fragments being higher than the longer fragments because there are more of them)  but all all just adding up to one sequence for every primer ( which itself may be annealing at the end, middle or any other part of a fragment where it finds its homology. Sequences are built up from the 3' end of the primer wherever it sticks.

Yes Paul, This is what I thought. So, how can I prove it's random? When I subject this cleaved product to restriction analysis I was expecting ladder as the cut may be random but I didn't get any ladder. The particular cut species population could drive this but I couldn't explain.

Can I clone these fragments? How to show this randomness? Can next gen sequencing help? I would appreciate some options I could try.Thanks for your valuable time. I really appreciate it.

if it is random I would expect that if you added too much enzyme and any pcr product or plasmid then given long enough a smear will be produced of random lengths and cut for even longer the smear will disappear as it all gets cut into tiny fragments. If non random then cutting for longer will make no difference and you will just get the same few/1 fragment..this is what you expect with a restriction enzyme. If you are expecting random fragments then you can digest and blunt end clone fragments and sequence with primers in the plasmid which should give you the ends of the sequences or even just cutting the cloned fragments out of the plasmid and showing that they are all different sizes will give you some idea of how random the process is. Cloning at least will separate the individual fragments and give you some clue to what process is happening

Yes, I will try cloning. Thanks for your input and time to help me out.