December 5, 1955

Dr. Alan Garen
Biophysical Leboratory
Purdve University
Lafayette, Indiana

Dear Alan:

Thank you for the recslealations cn mcterlal radiograchy. One trouble
is that ve won't be able to mke e specific correlation between P,. and gene
segregation: L.e., at the 4—cell stage, only one cell sill sogre rate out
viable recombinants, but wea can't tell which one. There are, however, any
number of reasons why the experiment is worth doing. Conceiwebly, even at
(say) the 32-cell stage, all the label will be in one cell: In that event, we
can sample parte of the clones (or subclones) for thoiy genctyne:, and look
for tf arong any of these left. Thit is, for the most useful ramite, this
experiment would be a refinement cf Cy's.

What woula you say to the idea cf doing this together-~ “ould Seymour be
likely to give you a few weeks off next summer or fall? You can tell him we'd
pay the teb. By then, I hope to have the techniaue to be able to do my end
of this kimi e6§ experiment, which I don't have now,

As to the diploids, we have never found Lp+/Lp+ derivatives from Lp+/Lp$
cells exposed toe lambda. ‘this is a puzzling result, but we want to do this
with Gal-transduction as a tag. That's one reason I'm struggling to make some
Upt/s Gai-/- diploids, but have had little luck so far. The other approach is
to use some marked lanbdes, and Esther has been hoping to cet this up soon.

The growth of T4r on Lp+/s may not be only a trivial experiment. Every
diploid population is contaminated with 5-10% segregants, so one will have to
do this on a statistical basis with single bursts, and guess whether the
fraction of bursts correspogds to the Lp® segregants or to the sum of these
with Lpt/s cells. With a good anti-T, serum, it might not be too difficult to
get a good count of infective centers, unless the heterozggotes show some un-
predicted and confusing behavior. --- I think perhaps one could also take advan-
tage of streptomycin resistance. Say we have a suspension with @5% Lp+/a;

10% ¢ and 5% + segregants, which is not too unreasonable. The same diploid is
segregating S™/S° s0 that most of the Lp® and all cf the +/s are sensitive to
streptomycin; the Lp? segregants are resistant. Infect with the T4, add serum
to inactivate free phage, and plate for infective centers with and whthout sm.
If the infective centers correspond to the TA~killed bacteria without sm, and
to the T4~killed Lp® with am, then both +/s and s are growing T4. If the infec-
tive centers correspond ohly to the killed Lp®, and are not reduced by am, then
only s grows the phage. I would not want to rely on a half—experiment without
such a check. Is the result worth the trouble? The systems where lambda
blocks Killing by other phages give easier answers. But I'll be glad to try

it if 1% seems important.

Sincerely,