APPLICATION FOR RESEARCH SUPPORT
To the Research Committee
Submitted by J. Lederberg, Department of Genetics

1950-1951

TITLE AND REASONS FOR STUDY

Bacterial Gemetics. The mechanism of genetic recombination
remains one of the crucial problems of bacterial genstics. The
controlled hybridization of bacteria requires the mest detailed
information from genetic and cytological studies on Escherichia
coli K-12, the bacterial strain in which gene recombination has
bean most clearly demonstrated.
PROG DURE

A, Cytological and genetic comparisons of haploid and diploid
cultures.

In a current project, exceptional diploid cultures of KE. coli
are being compared cytologically with the typical kaploid strains.
The diploids can be characterised as such by their genetic behavior,
namely that single cells give rise, during subsequent fissions,
to 'seeregant" cultures showing various combinations of the characters
of the ultimate parents. Cytologically, the diploids show a more
complex and disperse nuclear structure than the haploids. The observa-
tions may ultimately lead to the simplest, obvious interpretation:
that the nuclear units, or "chromosomes" are relatively redunlicated
in the diploid. However, these atudies mist be considerably extended
before any conclusions can be confirmed.

The primary difficulty, of course, lies in the extremely

small size of the bacteria nuclei, so that they can only barely be
resolved with the light microscope. To avoid subjective errors of
interpretation, especially since there is considerable variation

in nuclear morphology within haploid cultures, photomicrographic
records should be used as far as possible. Although Dr. Ris, of the
Department of Zoology, has cooperated very generously thusfar in
making available his facilities, no suitable photcmicrographic equip-
ment is available in the Department of Genetics, where it should

be located if any extensive use is to be made of it for this project.
It is requested, therefore, that support for this project be con-
tinued with a grant for a) renewal cf an assistantship for Miss \. i.
Lively, who is currently working on the cytological problem, and b)

microscopic and photomicrographie equipment.

B. Genetic mechanisms of bactericidal action.

The bactericidal action of radiations, and to some extent of
other agents, has been accounted for by many workers as due to the
induction of lethal mutations, like those which have been studied
in Drosophila (ef. Lea, Actions of Radiations on Living Cells). Al-
though uni-cellular organisms such as bacteria are especially suitable
for study of lethal radiological effects, it has not been possible
to rationalize the known genetic with the lethal effects of radiations,
bedause haploid cells are directly affected by lethal mutations, and
the killed cells are unavailable for further study. Lethal mutations
in diploid organisms should, however, have no immediate effect if they
are recessive in character, because the lethal mutation in one chromosome
will be protected by the unmutated homologue in the other chromosome.
However, lethal mutations can usually be detected by firther genetic
analysis. |

Surprisingly, preliminary experiments with ultra-violet light
have indicated an entirely distinct mechanism of action. Even with
highly lethal doses of UV, few or no recessive lethal mutations are
found. Nor can the results be accounted for by so-called dominant
}ethal mtations. Instead, the killing avpears to be accountable for
by effects on individual chromosomes or nuclei as units; that is,
diploid cells are made effectively haploid under the influence of UV.
Killing is possibly the succeeding stage in the series diploid-haploid-
death. such a sequence would alsc account for the sigmoid type of
dose-response curve found with UV.

ft is hoped to continue these studies, to ascertain the units
and kinetics of bactericidal action not only of UV but of Llonizing
radiations (X-rays, ganma rays), as well as of mutazenic chemicals
(e.g., nitrosen mustard), other disinfectants, and heat, No svecial
@quipaent ds required for this study, but request is made for a
second research arsistant to work on this and other voroblens,.

C. Uther problems.

The detailed study of the genetic behavior, in crosses, of
a number of different characters of %. coli K-12 is peing continued.
Until we have a clear picture of the genetic crzanization, in terms
of linkage maps and the mechanism of segregation, our understanding
of hybridization in this bacterium remains incomplete. There is strong
evidence for linear linkage, like that of higher forms of life, but
there are aiso anomiies not yet satisfactorily explained. This holds
in particular for the segregation ratios of diploid heterozyvotes,
which, according to the simpla model, should be 1: 1 for each character
pair, but which is as peturbed as 13: 1 in some instances. The only
approach to this perplexity is simply the sustained study of the linkage

behavior of a great many characters in crosses.
The validity of other projects in progress, for example, the genetic
control of bacterial ensymes, depends in part on the success of our
efforts at rectifying the gene systen.

A number of colleagues have felt that the scone of genetic recom-
bination, and presumably of sexuality in bacteria, should not be limited
to a single strain of a single species. Recombination is being looked
for, therefors, in other cultures, and finally has been demonstrated
in other cultures, first by L. Cavalli (Cambridge, Hngland}, and then
again here. Advantage is being taken of the inter-crossability of these
three bacteria (all 7. coli) to study the genetic basis of natur:.1 varietal

differencas,

FINANCIAL SUPPORT

Microscopic and micrographic equipment $ 550.

Research Assistants, 12 mo., [2} at 1150. 2300,

Hourly heip 200
TOTAL 3050.

Pp iN
Project Leader: Joshus cLederberz, Assistant Professor of
Gene tics.
Assistant : Hthelyn &. Lively, Graduate Student, began

residence Sept. 1949; Is pursuing her research
very satisfactorily

Assistant : To be selected from current applications.