Library construction -- mechanical shearing

[Shaun Tyler]

I=92m interested in sequencing a 40 kb plasmid (a native one not a clone)
for which I have a limited amount of material and no restriction map.=20
Since what I have was hard to come by I=92m hesitant to waste it in
establishing a restriction map in order to subclone more manageable
fragments.  Our sequencing capabilities are not a limiting factor so I
was thinking of using a shotgun approach with randomly sheared material
for a library and then closing the gaps by PCR and primer walking.=20
Unfortunately I have been unable to find any protocols for preparing
mechanically sheared DNA.  I know people occasionally use this method
for preparing genomic libraries but I=92m not sure how effective this
would be on something relatively small like a 40 kb plasmid.  Any input
would be greatly appreciated.


Shaun Tyler
DNA Core Facility
Laboratory Centre for Disease Control
Health Canada

Ph#:  (613) 941-6441
FAX#: (613) 957-1358

E-mail:  styler at hpb.hwc.ca