August 31, 1956

Dear Dr. Dienes:

Thank you for your letter of the 29th, and the om. I am looking for-
ward to mesting Dr. Sharp: would you ask him to call me at home (CEdar-3-
2968) when he arrives, or let us know where we can pick him up?

Since I wrote to you before, I have bean learning more and more, as mich
from reading and re-reading your papers as from new experiments. It is obvious
to me now that protoplasts are the initial large bodies, the starting point
of the L growth cycle. (I wish I had as clear < grasp of this when I wrote
that paper!)

I have also been able to achieve essentially q antitative yields: of
L colonies, both from E, coli K-12 and from Proteus 52;sckmendickkmex when
bacteria are inoculated into soft nutrient agar + penicillin + Mg + sucrose.
The sucrose is quite essential for this efficiency; it doubtless serves the
same function as your 3.5% NaCl, though I think more effectively. However,
I have had very limited success so far in propagating the L-colonies further
& still have to work this out. I have cause fo umierline lines 912, page
539 of your 1949 paper on Proteus! However, in my own faltering way, I om
learning some of your tricks——- hoping, of course, to be able to extend them
to my own favorite strains.

Unfortunately. I have not been able to obtain a floating growth even with
several trials on Proteus 52: it seams quite possible that the strain has
becoms less apt during storage here, and I would therefor: renev my re west
for the bacterial culture and flor a floating growth. It seems likely, as I
om gure you have in mind, that the floating growth is a mass which furnishes
itself the reiusiatte physic:] conditions for orcliferagdon.

I have madeca few observations cn slide cultures; these suggest that the
mode of proliferagion of the initial L colony is by budding of the protoplast.
Are there any direct microscopical observations of the role of minute gmnules
in proliferation? In retrospect, Klieneberger's observations are not entirely
convincing (I mean her filtration experiment). ven the cozrsesf filter removed
most of the visble count! The typical size of the reproductive units there may
have been very little smaller than that of the bacteria (which ure rather small
themselves).

Anyhow, there will be many things for Dr. Sharp and as to telk about; I
hope it will not be too long before I can pay my respects to you again in
person.

Yours sincerely,

Joshua Lederberg