Hi Folks,
One uses helper phage, R408, M13KO7 etc. with a phagemid to produce
packaged ss phagemid particles and also get the helper phage particles
by default. Does anyone know if you can take a helper phage and replace
the origin with say P15 (pACYC origin) to generate a plasmid with all
phage proteins minus the M13 origin. If so and one added a phagemid to
this system with a compatible origin say colE1, would one then get
solely packaged phagemid particles produced? If this was so simple it
would surely have been done by now so there must be a catch.
What I am trying to do is to preferentially package a phagemid with a
small insert but using a phagemid with two M13 origins in the same
direction (1). What happens with this system is that you get initiation
at one origin and termination at the other. DNA downstream of the second
origin is deleted so you land up just packaging the small insert DNA
between the two origins and say a colE1 origin and antibiotic selection
region after the second M13 origin gets deleted. You get an extremely
small packaged DNA fragment which is what I want. What I don't want is
the helper phage. I may be able to separate them by CsCl but again I may
not.
Duncan
(1) J.Mol.Biol. 153, 169-176 (1981)
--
The problem with being on the cutting edge is that you occasionally get
sliced from time to time....
Duncan Clark
DNAmp Ltd.
Tel: +44(0)1252376288
FAX: +44(0)8701640382
http://www.dnamp.comhttp://www.genesys.demon.co.uk
