G e N e t i c S , t e c H N o l o G Y , A N D S o c i e t Y 289 G e n e t i c s , t e c h n o l o G y , a n d s o c i e t y telomeres: the Key to immortality? Humans, like all multicellular organisms, grow old and die. As we age, our immune systems become less efficient, wound healing is impaired, and tissues lose re- silience. It has always been a mystery why we go through these age-related declines and why each species has a characteris- tic finite life span. Why do we grow old? Can we reverse this march to mortality? Some recent discoveries suggest that the answers to these questions may lie at the ends of our chromosomes. The study of human aging begins with a study of human cells growing in culture dishes. Like the organisms from which the cells are taken, cells in culture have a fi- nite life span. This replicative senescence was first noted by Leonard Hayflick in the 1960s. He reported that normal hu- man fibroblasts lose their ability to grow and divide after about 50 cell divisions. These senescent cells remain metaboli- cally active but can no longer prolifer- ate. Eventually, they die. Although we don’t know whether cellular senescence directly causes organismal aging, the evi- dence is suggestive. For example, cells de- rived from young people undergo more divisions than those from older people; cells from short-lived species stop grow- ing after fewer divisions than those from longer-lived species; and cells from pa- tients with premature aging syndromes undergo fewer divisions than those from normal patients. Another characteristic of aging cells involves their telomeres. In most mam- malian somatic cells, telomeres become shorter with each DNA replication be- cause DNA polymerase cannot synthe- size new DNA at the ends of each parent strand. However, as discussed in detail in this chapter, cells that undergo extensive proliferation, like embryonic cells, germ cells, and adult stem cells, maintain their telomere length by using telomerase—a re- markable RNA-containing enzyme that adds telomeric DNA sequences onto the ends of linear chromosomes. However, most somatic cells in adult organisms do not proliferate and do not contain active telomerase. Could we gain perpetual youth and vi- tality by increasing our telomere lengths? Studies suggest that it may be possible to reverse senescence by artificially increas- ing the amount of telomerase in our cells. When investigators introduced cloned telomerase genes into normal human cells in culture, telomeres lengthened, and the cells continued to grow past their typical senescence point. These studies suggest that some of the atrophy of tis- sues that accompanies old age could be reversed by activating telomerase genes. However, before we use telomerase to achieve immortality, we need to consider a potential serious side effect: cancer. Although normal cells shorten their telomeres and undergo senescence after a specific num ...

1. G e N e t i c S , t e c H N o l o G Y , A N D S o c i e t Y 289
G e n e t i c s , t e c h n o l o G y , a n d s o c i e t y
telomeres: the Key to immortality?
Humans, like all multicellular organisms, grow old and die. As
we age, our immune
systems become less efficient, wound
healing is impaired, and tissues lose re-
silience. It has always been a mystery why
we go through these age-related declines
and why each species has a characteris-
tic finite life span. Why do we grow old?
Can we reverse this march to mortality?
Some recent discoveries suggest that the
answers to these questions may lie at the
ends of our chromosomes.
The study of human aging begins with
a study of human cells growing in culture
dishes. Like the organisms from which the
cells are taken, cells in culture have a fi-
nite life span. This replicative senescence
was first noted by Leonard Hayflick in
the 1960s. He reported that normal hu-
man fibroblasts lose their ability to grow
and divide after about 50 cell divisions.
These senescent cells remain metaboli-
cally active but can no longer prolifer-
ate. Eventually, they die. Although we
don’t know whether cellular senescence

2. directly causes organismal aging, the evi-
dence is suggestive. For example, cells de-
rived from young people undergo more
divisions than those from older people;
cells from short-lived species stop grow-
ing after fewer divisions than those from
longer-lived species; and cells from pa-
tients with premature aging syndromes
undergo fewer divisions than those from
normal patients.
Another characteristic of aging cells
involves their telomeres. In most mam-
malian somatic cells, telomeres become
shorter with each DNA replication be-
cause DNA polymerase cannot synthe-
size new DNA at the ends of each parent
strand. However, as discussed in detail in
this chapter, cells that undergo extensive
proliferation, like embryonic cells, germ
cells, and adult stem cells, maintain their
telomere length by using telomerase—a re-
markable RNA-containing enzyme that
adds telomeric DNA sequences onto the
ends of linear chromosomes. However,
most somatic cells in adult organisms do
not proliferate and do not contain active
telomerase.
Could we gain perpetual youth and vi-
tality by increasing our telomere lengths?
Studies suggest that it may be possible to
reverse senescence by artificially increas-
ing the amount of telomerase in our cells.
When investigators introduced cloned
telomerase genes into normal human

3. cells in culture, telomeres lengthened,
and the cells continued to grow past their
typical senescence point. These studies
suggest that some of the atrophy of tis-
sues that accompanies old age could be
reversed by activating telomerase genes.
However, before we use telomerase to
achieve immortality, we need to consider
a potential serious side effect: cancer.
Although normal cells shorten their
telomeres and undergo senescence after
a specific number of cell divisions, can-
cer cells do not. More than 80 percent
of human tumor cells contain telomerase
activity, maintain telomeres, and achieve
immortality. Those that do not contain
active telomerase use a less well under-
stood mechanism known as ALT (for “al-
ternative lengthening of telomeres”).
These observations have motivated
scientists to devise new cancer therapies
based on the idea that agents that inhibit
telomerase might destroy cancer cells by
allowing telomeres to shorten, thereby
forcing the cells into senescence. Because
most normal human cells do not express
telomerase, such a therapy might target
tumor cells and be less toxic than most
current anticancer drugs. Many such anti-
telomerase drugs are currently under de-
velopment, and some are in clinical trials.
Will a deeper understanding of telo-
meres allow us to both arrest cancers and

4. reverse the descent into old age? Time
will tell.
Your Turn
Take time, individually or in groups, to answer the following
questions. Investigate the references and links
to help you understand some of the re-
search on telomeres, aging, and cancer.
1. How might our knowledge about
telomeres and telomerase be
applied to anti-aging strategies? Are
such strategies or therapies being
developed?
Sources of information can be obtained by us-
ing the PubMed Web site (http://www.ncbi.
nlm.nih.gov/sites/ entrez?db=pubmed).
2. One anti-telomerase drug, called
GRN163L, is being developed by
Geron Corporation as a treatment
for cancer. How does GRN163L
work? What is the current status of
GRN163L clinical trials? What are
some possible side effects for anti-
telomerase drugs?
Read about this drug and its clinical tri-
als on the Geron Web site at http://www.
geron.com. Search on PubMed for scientific
papers dealing with GRN163L’s anticancer
effects.

5. 3. People suffering from chronic stress
appear to have more health prob-
lems and to age prematurely. Is there
any evidence that chronic stress,
poor health, and telomere length are
linked? How might stress affect telo-
mere length or vice versa?
Some recent papers suggest how these phe-
nomena may be linked. One such paper is Epel,
E. S., et al. 2004. Accelerated telomere
shortening in response to life stress.
Proc. Natl. Acad. Sci. USA 101(49): 17312–
17315.
4. In 2006, the Lasker Award for Ba-
sic Medical Research was awarded
to Drs. Elizabeth Blackburn, Carol
Greider, and Jack Szostak, who subse-
quently were awarded the 2009 Nobel
Prize in Physiology or Medicine. How
did the intersections of people, ideas,
and good fortune lead to their discov-
ery of telomerase and its role in aging
and cancer? What is the future for this
research?
Listen to interviews with these scientists,
in which they tell their stories about their
research and where they see the field
going, at http://www.laskerfoundation.
org/ 2006videoawards.
KLUG72412_10_SE_C11_pp269-293.indd 289
7/6/11 6:19 AM

6. 3rd Proof
6A This week, you are learning about inductive reasoning and
deductive reasoning. Inductive reasoning moves from specific
instances to a general conclusion while deductive reasoning is
the opposite--it moves from a true generalizable principle
known to be true to a specific conclusion. For this forum,
provide an example of inductive reasoning and an example of
deductive reasoning. Do you feel most research uses deductive
reasoning or inductive reasoning?
Peer Responses: Students are required to respond to at least two
(2) other student’s initial postings (and the instructor) with
significant comments that have substance. Peer responses are
due no later than 11:55 p.m. ET on Sunday, Day 7 of each week.
All peer responses must be substantial and significant and
should be no less than 100 words
RESPOND TO STUDENTS
WARSH
Good morning all,
After reading through the lessons I was able to grasp a much
better understanding of inductive and deductive research. I
realized when I was gaining a better understanding of what
inductive and deductive research techniques are that, I was
generally using inductive or deductive research techniques any
time I was doing research without even noticing it. According to
Chapman (2009), “deduction is a form of reasoning in which
one proceeds from general principles or laws to specific cases.
Induction is a form of reasoning in which one arrives at a
general principles or laws by generalizing over specific cases”
(p.48).
The best description I found that helped me understand
deductive reasoning was; A = B, and B=C, therefore A=C. A
valid example of this would be, Labradors are dogs, dogs are
animals, and therefore Labradors are animals.

7. Inductive reasoning is usually used when very little existing
knowledge is available. Inductive reasoning generally has three
steps; an observation, observe a pattern, develop a theory. AN
example on inductive reasoning would be, a low-cost hotel is
booked, another 5 low-cost hotel rooms are booked, and
therefore low-cost hotels are always booked.
Inductive and deductive reasoning can both be used in research
studies, I think it depends on what you’re trying to discover. If
you’re trying to test hypothesis and theories through
experimentation then you would want to use a deductive
approach. If you have a topic of interest with little known
information or data you should use an inductive approach to
form a hypothesis or theory.
Reference:
Chapman, S., & Routledge, C. (2009). Induction/Deduction.
Key ideas in linguistics and the philosophy of language.
Edinburgh University Press.
JJ
Induction is described as generalizing a conclusion from
a single or more phenomenon based on specific observation,
whereas deduction is described as applying reasoning to general
theories and principles to come to a specific conclusion (Meng,
n.d.).
The best description I’ve heard to differentiate inductive
reasoning and deductive reasoning is that induction starts at the
bottom and works its way up, whereas deduction starts at the
top and works its way down. In the many versions of Sherlock
Holmes, his methods of case solving are often described as
deduction, but in most portrayals his methods of case solving
are done with induction.
An example of deductive reasoning would be a mathematical
proof, such as the Pythagorean Theorum. An example of
inductive reasoning would be I’ve met several people from
England, they all liked tea, so I conclude that everyone from
England likes tea.
References

8. Meng, H. (n.d.). The relationship between reduction and
deductive & inductive reasoning. Retrieved
from https://pdfs.semanticscholar.org/fa34/2b77370c6743519c2
432f13654dd952525ac.pdf
6B RESPOND TO PEER
Peer Responses: Students are required to respond to at least two
(2) other student’s initial postings (and the instructor) with
significant comments that have substance. Peer responses are
due no later than 11:55 p.m. ET on Sunday, Day 7 of each week.
All peer responses must be substantial and significant and
should be no less than 150 words
WARS
The scientific peer-reviewed article I chose that focuses on our
assigned topic; find an article on sports nutrition to discover
what an athlete eats is critical for their success is titled,
“Influence of nutrition and genetics on performance: a pilot
study”. A lot of factors can affect ones day to day performance,
some example could be: sleep, genetics, stress, mental alertness,
or nutrition. The study I picked focuses on nutrition and
genetics and how they play a factor in energy levels during a
gymnastics practice. This research study was designed to survey
17 girls aged between the ages of 10-14. Before one practice the
girls were given a carbohydrate based snack before and after the
practice. On a different day they were given a protein based
snack before and after the practice. The study concluded that
the girls had more energy on the days where they ate a
carbohydrate heavy snack than the days when they ate a protein
based snack. The girls claimed that in addition to having more
energy they also said their performance improved after eating a
carbohydrate snack versus when they ate a protein snack.
I do not think the study can draw a definitive conclusion. I
believe that the sample size was way too small and only used
one sport. For the study to draw a better conclusion it should
have: used both male and females, had more athletes studied,
used different areas of the country, used different types of

9. carbohydrate and protein based snacks, and the study should
have been over the course of a few months. The study needs to
be longer due to the multiple factors that I mentioned above
resulting in day to day performance. If you did not get much
sleep the night before training, than that will play a major effect
on your performance, it would not matter whether you eat a
carbohydrate or protein based snack.
I believe that nutrition does play a major role in sports success.
I tend to eat clean Monday through Saturday and splurge and eat
unhealthier options on Sunday. I play soccer on Monday nights
and Thursday nights, and I always have less energy on Monday
nights during the game than I do on Thursday nights and I
believe it is because I eat a lot worse on Sunday. Whereas by
Thursday night I have four consecutive days of eating healthy
leading up to my game and I always feel faster, and have better
performances.
References:
Amato, A., Contro, V., Galassi, C., Proia. P., Macchiarella, A.,
Sabatino, E., & Sacco, A. (2017). Influence of nutrition and
genetics on performance: a pilot study. Human Movement. Vol
18, p, 12-16.
DECK
My selected source for this week’s forum is titled “Timing,
Optimal Dose and Intake Duration of Dietary Supplements with
Evidence-Based Use in Sports Nutrition” from the Journal of
Exercise Nutrition and Biochemistry.
In this study, researchers looked at optimal timing for main
dietary supplements such as beta-alanine, nitrate, caffeine,
creatine, sodium bicarbonate, carbohydrates, and protein. Some
of the findings were very interesting such as “abstention from
caffeinated foods and beverages for a few days prior to an event
may maximize performance effects. Recently, however, morning
ingestion of caffeine improved neuromuscular function
(contraction velocity.)” (Naderi et al., 2016) Researches also
discovered that 3-6 mg/kg of bodyweight taken 30-60 minutes

10. before exercises increases time until exhaustion, improved work
capacity, and reduced perception of effort during exercise.
Creatine loading, or taking in 3 – 5 g of protein a day for
several weeks, has been shown to increase levels. Excess levels
can enhance glycogen storage, increase the capacity of the
phosphagen energy system, and play a role in gene
transcription. However, timing is the most important component
to reap the benefits. The first study on creatine that was
conducted found that post-exercise creatine ingestion is far
more beneficial in comparison to pre-workout creatine. Similar
results were discovered for a majority of the different
supplements.
The conclusion of this study was that “Several nutritional sports
foods and supplements are effective at influencing energy
supply, with substantial evidence for carbohydrate and creatine
supplements and for physiological buffering agents such as β-
alanine and sodium bicarbonate.” (Naderi et al. 2016) I think
that the study made appropriate conclusions from the data, and
that it was designed to correctly to address the hypothesis.
I think that this article was very informed, and presented a wide
variety of information that many more people should use. One
of the most important ones that was covered was the protein
window, and showing that there is no “30 minute anabolic
window” after the gym, where either you get protein in or lose
your hard work. The study showed that it was actually 16-48
hours, and that Caesin right before sleep actually helped with
recovery more while the person was sleeping. Overall I think it
was a very useful and interesting article that more people
should take the time to read if they work out or train in any
capacity.
Reference:
Naderi, A., de Oliveira, E., Ziegenfuss, T., Willems, M., &
Naderi, A. (2016). Timing, Optimal Dose and Intake Duration
of Dietary Supplements with Evidence-Based Use in Sports
Nutrition. Journal of Exercise Nutrition & Biochemistry, 20(4),

11. 1–12. https://doi.org/10.20463/jenb.2016.0031
Name: Date:
Critical Reading Form for Meselson & Stahl – 10 points
Type in answers – Save & Upload to Canvas
Purpose:In 1-2 sentences, what was the research question the
authors were trying to answer and why?
Central Thesis:Using only 1-2 sentences, what was the authors’
thesis, or hypothesis?
Key Claims:What are two statements that are used to support
the thesis? Conclusions or discussion.
1.
2.
Evidence:What evidence do the author’s use to support each of
the key claims you identified?
These are results of experiments.
1.
2.
Figure Analysis (Figure 4 – Meselson & Stahl)
What is shown in part a?

12. What is shown in part b?
What do the numbers under the generation heading mean?
Why is the pattern different between generation 0
and generation 2.5?
Why is the pattern different between generation 0
and generation 4.1?
671
Appearances of regeneration from naked masses of protoplasm
have sometimes been noted, but never in examples in which it
had been established that there were no small, fully formed cells
from which the growth might have arisen.
It is, of course, possible that cultural conditions will be found
under which masses of naked protoplasm will revert to cellular
growth, such as has been reported by Ne&as4 in Saccharomyces
cerevisiae and by Pease"' in Proteus vulgaris.
1 C. Weibull, Symposium Soc. Gen. Microbiol., 6, 111, 1956.
2J. Lederberg, these PROCEEDINGS, 42, 574, 1956; K.
McQuillen, Biochim. et Biophys. Acta, 27,
410,1958.
3 K. McQuillen, Symposium Soc. Gen. Microbiol, 6, 127, 1956;
S. Spiegelman, in The Chemical Basis of Heredity, ed. W. D.
McElroy and B. Glass (Baltimore: Johns Hopkins Press, 1957),
pp. 232-267; J. Spizizen, these PROCEEDINGS, 43, 694, 1957;
D. Fraser, H. R. Mahler, A. L. Shug, and C. A. Thomas, Jr.,
these PROCEEDINGS, 43, 939, 1957; E. Chargaff, H. M.
Schulman, and H. S. Shapiro, Nature, 180, 151, 1957.
40. Necas, Biol. Zentr., 75, 268, 1956; A. A. Eddy and D. H.

13. Williamson, Nature, 179, 1252, 1957.
5 Mary R. Emerson, in manuscript. Our mutant strain, Em
11200, has been shown to be allelic to a number of others of
independent origin: B 135 and M 16 obtained from Dr. D. D.
Perkins, Stanford University; "wooly," from Dr. H. L. K.
Whitehouse, Cambridge University; and "ginger" and an
unnamed mutant from Dr. J. R. S. Fincham, University of
Leicester. From published descriptions it is likely that these are
also allelic to the "cut" mutant of Dr. H. Kuwana
(Cytologia, 18,235, 1953).
6 Mary B. Mitchell and H. K. Mitchell, these PROCEEDINGS,
38, 442, 1952.
Mary B. Mitchell, H. K. Mitchell, and A. Tissieres, these
PROCEEDINGS, 39, 606, 1953. 8 G. W. Beadle and E. L.
Tatum, these PROCEEDINGS, 27, 499, 1941. 9 The
hemicellulase preparation was obtained from the Nutritional
Biochemical Corporation, Cleveland 28, Ohio, who inform us
that it is a concentrate of microbiological origin, containing
cellulase, gumase, and maltase in addition to hemicellulase. A
single test in our laboratory indi cated strong chitinase activity.
1OJ. Lederberg and Jacqueline St. Clair, J. Bacteriol., 75, 143,
1958. 11 Phyllis Pease, J. Gen. Microbiol., 17, 64, 1957.
THE REPLICATION OF DNA IN ESCHERICHIA COLI* BY
MATTHEW MESELSON AND FRANKLIN W. STAHL
GATES AND CRELLIN LABORATORIES OF CHEMISTRY, t
AND NORMAN W. CHURCH LABORATORY OF CHEMICAL
BIOLOGY, CALIFORNIA INSTITUTE OF TECHNOLOGY,
PASADENA, CALIFORNIA
Communicated by Max Delbrick, May 14, 1958
Introduction.-Studies of bacterial transformation and
bacteriaphage infection'-'
strongly indicate that deoxyribonucleic acid (DNA) can carry
and transmit heredi tary information and can direct its own
replication.
Hypotheses for the mechanism of DNA replication differ in the
predictions they make concerning the distribution among

14. progeny molecules of atoms derived from parental molecules.6
Radioisotopic labels have been employed in experiments
bearing on the distribu tion of parental atoms among progeny
molecules in several organisms.6-9 We anticipated that a label
which imparts to the DNA molecule an increased density might
permit an analysis of this distribution by sedimentation
techniques.
To this end, a method was developed for the detection of small
density differences among
6;72
bacterial Distance fuged stages FIG. at 1.-Ultraviolet in lysate
31,410 from the banding containing the rpm axis in absorption a
of of approximately CsCl DNA rotation solution photographs
from increases E. as 108 described coli. lysed showing toward
cells An in was aliquot the successive the centri- right. text. of
after The number reaching beside 31,410 rpm. each photograph
gives the time elapsed
macromolecules.'0 By use of this method, we have observed the
distribution of the heavy nitrogen isotope N15 among molecules
of DNA following the transfer of
a uniformly N"5-labeled, exponentially growing bacterial
population to a growth medium containing the ordinary nitrogen
isotope N'4.
Density-Gradient Centrifugation.- A small amount of DNA in a
concentrated solution of cesium chloride is centrifuged until
equilibrium is closely approached.
BIOLOGY: MESELSON AND STAHL
673
The opposing processes of sedimentation and diffusion have
then produced a stable concentration gradient of the cesium
chloride. The concentration and pressure gradients result in a
continuous increase of density along the direction
of centrifugal force.
The macromolecules of DNA present in this density gradient are
driven by the centrifugal field into the region where the solution

15. density is equal to their own buoyant density.'
This concentrating tendency is opposed by diffusion, with the
result that at equilibrium a single species of DNA is distributed
over a band whose width is inversely related to the molecular
weight of that species
(Fig. 1).
If several different density species of DNA are present, each
will form a band at the position where the density of the CsCl
solution is equal to the buoyant density In this way DNA
labeled with heavy nitrogen (N15) may be of that species.
a
N'14 N5
b
A two at CsCl mixture FIG. 44,770 bands solution 2-a: rpm. of
of N'4 Fig. as The b: gin. described and 2a. A resolution cm.
N"1 microdensitometer The bacterial -- in separation the of text.
N14 lysates, DNA between The tracing each photograph from
containing the showing N"1 peaks DNA was the about
corresponds taken DNA by 10' density-gradtient after
distribution lysed to 24 cells, a hounrs difference was in of the
centrifugation. centrifugation centrifuged region in buoyant of
the in density of 0.014
Figure 2 shows the two bands formed as a result of centrifuging
a mixture of approximately equal amounts of N14 and N1I-
Escherichia coli DNA.
resolved from unlabeled DNA.
In this paper reference will be made to the apparent molecular
weight of DNA
samples determined by means of density-gradient
centrifugation. A discussion has been given"' of the
considerations upon which such determinations are based,
as well as of several possible sources of error.'2
Experimental.-Escherichia coli B was grown at 360 C. with
aeration in a glucose salts medium containing ammonium
chloride as the sole nitrogen source." The growth of the

16. bacteria) population was followed by microscopic cell counts
and by colony assays (Fig. 3).
Bacteria uniformly labeled with N"5 were prepared by growing
washed cells for
S.674 BIOLOGY: MESELSON AND STAHL PROC. N. A.
14 generations (to a titer of 2 X 108/ml) in medium containing
100 ,ug/ml of N'5H4Cl An abrupt change to N14 medium was
then ac of 96.5 per cent isotopic purity. complished by adding
to the growing culture a tenfold excess of N14H4Cl, along with
ribosides of adenine and uracil in experiment 1 and ribosides of
adenine. guanine, uracil, and cytosine in experiment 2, to give a
concentration of 10lg/ml During subsequent growth the
bacterial titer was kept between of each riboside.
-14 -12 -10 -8 -6 -4 -2 0 TIME (hours)
2 4 6 8
medium. withdrawals tion, Thereafter, centrifugation, on FIG.
the the ordinates 3.-Growth generation The during and values
the give additions. of the actual time bacterial the on period the
was actual titer During ordinates 0.81 populations when was
titers hours the kept samples of during period in the first
between Experiment cultures were this in of N sampling later 15
being 1 and and up period 1 then and withdrawn to 2 for the X
in 0.85 density-gradient have N14 108 time hours been medium.
by for of additions in corrected density-gradient addition
Experiment The centrifuga- of of for values fresh N'4. the 2.
1 and 2 X 108/ml by appropriate additions of fresh N 14
medium containing ribosides. Samples containing about 4 X 109
bacteria were withdrawn from the culture just before the
addition of N14 and afterward at intervals for several
generations. Each sample was immediately chilled and
centrifuged in the cold for 5 minutes at After resuspension in
0.40 ml. of a cold solution 0.01 M in NaCl and 1,800 X g.
0.01 M in ethylenediaminetetra-acetate (EDTA) at pH 6, the
cells were lysed by the
addition of 0.10 ml. of 15 per cent sodium dodecyl sulfate and

17. stored in the cold.
2
2
gradient of cess on CsCl FIG. centrifugation each of solution
Ni4 4-a: centrifugation substrates increases Ultraviolet at
44,770 to to of a growing the lysates absorption rpm under of
Ni"labeled bacteria photographs the conditions sampled culture.
showing at described Each various DNA photograph times in
bands the after text. was resulting the taken The addition
density from after of 20 density- position Ni4 of an hours the
ex- in units the the and DNA species measurements base
unlabeled of bands of the photograph. line DNA generation
shown is DNA directly corresponds of in shown bacterial the
The time. proportional adjacent in time to the right. The growth
the of lowermost photographs. relative generation sampling
Regions to presented the position frame, concentration is of
times measured equal in The which Fig. of for density its
microdensitometer Experiments 3. density serves band When
from of 6 DNA. occupy between Microdensitometer the as
allowance is just a time density 1 the The half-labeled and the
of same pen is degree 2 the bands made reference. were
displacement horizontal addition of estimated of for tracings is
labeling provided fully the A of relative labeled test of above
from of the by of a the conclusion that the DNA in the band of
intermediate the amounts at50 ±[ frame 2 per of showing cent
DNA of in the the the mixture distance three peaks, of between
generations the the peak N" 0 of and and intermediate 1.9. NI,'
peaks. density is found to be centered
BIOLOGY: MESELSON AND STAHL
PROC. NT, A. S.
For density-gradient centrifugation, 0.010 mnl. of the dodecyl
sulfate lysate was added to 0.70 ml. of CsCl solution buffered at
pH 8.5 with 0.01 M tris(hydroxy methyl)aminomethane. The

18. density of the resulting solution was 1.71 gm. cm.-3 This was
centrifuged at 140,000X g. (44,770 rpm) in a Spinco model E
ultracentri fuge at 250 for 20 hours, at which time the DNA had
essentially attained sedimenta tion equilibrium. Bands of DNA
were then found in the region of density 1.71 gm. cm.-3, well
isolated from all other macromolecular components of the
bacterial lysate. Ultraviolet absorption photographs taken
during the course of each cen trifugation were scanned with a
recording microdensitometer (Fig. 4). The buoyant density of a
DNA molecule may be expected to vary directly with The
density gradient is constant in the the fraction of N15 label it
contains. region between fully labeled and unlabeled DNA
bands. Therefore, the degree of labeling of a partially labeled
species of DNA may be determined directly from the relative
position of its band between the band of fully labeled DNA and
the The error in this procedure for the determination of band of
unlabeled DNA. the degree of labeling is estimated to be about
2 per cent. Results.-Figure 4 shows the results of density-
gradient centrifugation of lysates of bacteria sampled at various
times after the addition of an excess of N'4-containing
substrates to a growing N1"-labeled culture. It may be seen in
Figure 4 that, until one generation time has elapsed, half labeled
molecules accumulate, while fully labeled DNA is depleted.
One generation time after the addition of N 14, these half-
labeled or "hybrid" molecules alone are observed. Subsequently,
only half-labeled DNA and completely unlabeled DNA
When two generation times have elapsed after the addition of
N'4, half-labeled and unlabeled DNA are present in equal
amounts.
are found.
Discussion.-These results permit the following conclusions to
be drawn regard ing DNA replication under the conditions of
the present experiment.
1. The nitrogen of a DNA molecule is divided equally between
two subunits which remain intact through many generations.
The observation that parental nitrogen is found only in half-

19. labeled molecules at all times after the passage of one
generation time demonstrates the existence in each DNA
molecule of two subunits containing equal amounts of nitrogen.
The finding that at the second generation half-labeled and
unlabeled molecules are found in equal amounts shows that the
number of surviving parental subunits is twice the number of
parent molecules initially present. That is, the subunits are
conserved. 2. Followinq replication, each daughter molecule has
received one parental subunit. The finding that all DNA
molecules are half-labeled one generation time after the
addition of N'4 shows that each daughter molecule receives one
parental sub unit.'4 If the parental subunits had segregated in
any other way among the daughter molecules, there would have
been found at the first generation some fully labeled and some
unlabeled DNA molecules, representing those daughters which
received two or no parental subunits, respectively. 3. The
replicative act results in a molecular doubling. This statement is
a corollary of conclusions 1 and 2 above, according to which
each parent molecule passes on two subunits to progeny
molecules and each progeny
BIOLOGY: MESELSON AND STAHL
molecule receives just one parental subunit.
677
It follows that each single molecular reproductive act results in
a doubling of the number of molecules entering into that act.
The above conclusions are represented schematically in Figure
5. The Watson-Crick Model.-A molecular structure for DNA has
been proposed by Watson and Crick.15 It has undergone
preliminary refinement'6 without alteration of its main features
and is supported by physical and chemical studies."7 The
structure consists of two polynucleotide chains wound helically
about a common axis. The nitrogen base (adenine, guanine,
thymine, or cytosine) at each level
ORIGINAL 7
PARENT MOLECULE

20. tions. equally receives the FIG. data 5.-Schematic between one
presented of these. two in subunits. representation Fig. The 4.
subunits The Following nitrogen of are the conserved
duplication, conclusions of each through DNA each drawn
molecule successive daughter in the is text molecule duplica-
divided from
on one chain is hydrogen-bonded to the base at the same level
on the other chain. Structural requirements allow the occurrence
of only the hydrogen-bonded base pairs adenine-thymine and
guanine-cytosine, resulting in a detailed complemen tariness
between the two chains.
This suggested to Watson and Crick"8 a definite and
structurally plausible hypothesis for the duplication of the DNA
molecule. According to this idea, the two chains separate,
exposing the hydrogen-bonding sites of the bases.
Then, in accord with the base-pairing restrictions, each chain
serves as a template for the synthesis of its complement.
Accordingly, each daughter molecule contains one of the
parental chains paired with a newly synthesized chain (Fig. 6).
BIOLOGY: MESELSON AND STAHL
The results of the present experiment are in exact accord with
the expectations of the Watson-Crick model for DNA
duplication. However, it must be emphasized that it has not
been shown that the molecular subunits found in the present ex
periment are single polynucleotide chains or even that the DNA
molecules studied here correspond to single DNA molecules
possessing the structure proposed by Watson and Crick.
However, some information has been obtained about the
molecules and their subunits; it is summarized below.
parent proposed two chain tains FIG. molecules one (white).
chains 6.-Illustration by of Watson the remain each Upon
parental with and intact, continued of one Crick. the chains
parental so mechanism that Each duplication, (black) there

21. chain. daughter paired will of DNA the always with molecule
two duplication be one original found con- new
The DNA molecules derived from E. coli by detergent-induced
lysis have a buoyant density in CsCl of 1.71 gm. cm.-3, in the
region of densities found for T2 and T4 bacteriophage DNA,
and for purified calf-thymus and salmon-sperm DNA. A highly
viscous and elastic solution of N14 DNA was prepared from a
dodecyl sulfate lysate of E. coli by the method of Simmons19
followed by deproteinization with chloroform. Further
purification was accomplished by two cycles of preparative
density-gradient centrifugation in CsCl solution. This purified
bacterial DNA was found to have the same buoyant density and
apparent molecular weight, 7 X 106, as the DNA of the whole
bacterial lysates (Figs. 7, 8).
BIOLOGY: MESELSON AND STAHL
Heat Denaturation.-It has been found that DNA from E. coli
differs importantly from purified salmon-sperm DNA in its
behavior upon heat denaturation. Exposure to elevated
temperatures is known to bring about an abrupt collapse of the
relatively rigid and extended native DNA molecule and to make
available for acid-base titration a large fraction of the
functional groups presumed to be blocked by hydrogen-bond
formation in the native structure. 19, 20, 21, 22
Rice and Doty22 have reported that this collapse is not
accompanied by a reduction in molecu
These findings are corroborated by lar weight as determined
from light-scattering.
density-gradient centrifugation of salmon-sperm DNA.23 When
this material is
relative of FIG. Fig. 8.-The 7 concentration plotted square
against of of the DNA. the width logarithm The of the divisions
of band the persity the band. the to along any DNA the absence
point the located weight of Linearity abscissa of the of such at
density banded average the of set a corresponding plot this off

22. heterogeneity, DNA. molecular intervals plot is directly
indicates The of position weight value proportional 1 the mm.2.
monodis- slope of of in the the the at In
FIG. 7.-Microdensitometer tracing of an ultraviolet optical
density absorption in the region photograph of a band showing
of N14 the E. coli DNA at equilibrium. About 2 fg. of DNA
purified at 31,410 as rpm described at 250 in in 7.75 the molal
text was CsCl centrifuged atpH 8.4. The over the density region
gradient of the maximum band is and essentially is indicates
0.057 gm./cm.4. constant a the The ant optical density position
density of of 1.71 the the above concentration gm. cm.-' the
base In of line this DNA is directly tracing buoy- in the
proportional rotating DNA at the centrifuge maximum to cell. is
about The 50 concentration ,g./ml. of the weight responding
sodium slope for corresponds the to salt. a Cs-DNA molecular
to an salt weight apparent of 9.4 of 7.1 X molecular X 10., 10.
cor- for
kept at 1000 for 30 minutes either under the conditions
employed by Rice and Doty or in the CsCl centrifuging medium,
there results a density increase of 0.014 gm. cm.r3 with no
change in apparent molecular weight. The same results are ob
tained if the salmon-sperm DNA is pre-treated at pH 6 with
EDTA and sodium
Along with the density increase, heating brings about a sharp
dodecyl sulfate.
reduction in the time required for band formation in the CsCl
gradient.
In the absence of an increase in molecular weight, the decrease
in banding time must be ascribed10 to an increase in the
diffusion coefficient, indicating an extensive col lapse of the
native structure.
680
BIOLOGY: MESELSON AND STAHL
hours the denaturation. unheated densitometry for density FIG.
position comparison. of 9.-The has centrifugation N1" been

23. indicated. -3 bacterial of Each and dissociation removed an
Heating a ultraviolet smooth reduction lysates. in Unheated by
CsCi has of subtraction. curve the brought of absorption
solution Heated about lysate subunits connects lysate about half
at was A: photograph 44,770 of points in added A E. alone the a
mixture coli density rpm. apparent obtained to gives DNA this
taken of The one increase experiment heated upon molecular by
after baseline band for micro- heat and one 20 in of weight
generation brid The 0.016 in Fig. density DNA gm. 4. of cm. C:
the contained in difference A N14 DNA. mixture growth in is B:
this of 0.015 medium. Heated heated lysate gm. N14 lysate cm.
forms Before and -' of only heated N16 heat one bacteria
denaturation, N" band, bacterial as grown may lysates. the be
seen hy
The decrease in banding time and a density increase close to
that found upon heating salmon-sperm DNA are observed (Fig.
9, A) when a bacterial lysate containing uniformly labeled N"5
or N14 E. coli DNA is kept at 100° C. for 30 minutes in the
CsCl centrifuging medium; but the apparent molecular weight of
681
the heated bacterial DNA is reduced to approximately half that
of the unheated material. Half-labeled DNA contained in a
detergent lysate of N'5 E. coli cells grown for one generation in
N14 medium was heated at 1000 C. for 30 minutes in the CsCl
centri This treatment results in the loss of the original half-
labeled fuging medium. material and in the appearance in equal
amounts of two new density species, each with approximately
half the initial apparent molecular weight (Fig. 9, B). The
density difference between the two species is 0.015 gm. cm.-,
close to the increment produced by the N'6 labeling of the
unheated DNA. This behavior suggests that heating the hybrid
molecule brings about the dis sociation of the NI5-containing
subunit from the N'4 subunit. This possibility was tested by a
density-gradient examination of a mixture of heated N'5 DNA
and heated N14 DNA (Fig. 9, C). The close resemblance
between the products of heating hybrid DNA (Fig. 9 B) and the

24. mixture of-products obtained from heating N14 and N'5 DNA
separately (Fig. 9, C) leads to the conclusion that the two
molecular subunits have indeed dissociated upon heating. Since
the apparent molecular weight of the subunits so obtained is
found to be close to half that of the intact molecule, it may be
further concluded that the subunits of the DNA molecule which
are conserved at duplication are single, continuous structures.
The scheme for DNA duplication proposed by Delbrfick24 is
thereby ruled out. To recapitulate, both salmon-sperm and E.
coli DNA heated under similar conditions collapse and undergo
a similar density increase, but the salmon DNA retains its initial
molecular weight, while the bacterial DNA dissociates into the
two subunits which are conserved during duplication. These
findings allow two On the one hand, if we assume that salmon
DNA con different interpretations. tains subunits analogous to
those found in E. coli DNA, then we must suppose that the
subunits of salmon DNA are bound together more tightly than
those of the On the other hand, if we assume that the molecules
of salmon DNA bacterial DNA. do not contain these subunits,
then we must concede that the bacterial DNA molecule is a
more complex structure than is the molecule of salmon DNA.
The latter interpretation challenges the sufficiency of the
Watson-Crick DNA model to explain the observed distribution
of parental nitrogen atoms among progeny molecules.
Conclusion.-The structure for DNA proposed by Watson and
Crick brought forth a number of proposals as to how such a
molecule might replicate. proposals6 make specific predictions
concerning the distribution of parental atoms The results
presented here give a detailed answer to among progeny
molecules. the question of this distribution and simultaneously
direct our attention to other problems whose solution must be
the next step in progress toward a complete understanding of
the molecular basis of DNA duplication. What are the molecular
structures of the subunits of E. coli DNA which are passed on
intact to each daughter molecule? What is the relationship of
these subunits to each other in a DNA molecule? What is the

25. mechanism of the synthesis and dissociation of the sub units in
vivo? Summary.-By means of density-gradient centrifugation,
we have observed the distribution of N'5 among molecules of
bacterial DNA following the transfer of a uniformly N'5-
substituted exponentially growing E. coli population to N'4
medium.
These
PROC. N. A. S.
We find that the nitrogen of a DNA molecule is divided equally
between two physi cally continuous subunits; that, following
duplication, each daughter molecule receives one of these; and
that the subunits are conserved through many duplica
tions.
* Aided by grants from the National Foundation for Infantile
Paralysis and the National In stitutes of Health.
t Contribution No. 2344.
R. D. Hotchkiss, in The Nucleic Acids, ed. E. Chargaff and J.
N. Davidson (New York:
· Academic Press, 1955), p. 435; and in Enzymes: Units of
Biological Structure and Function, ed.
· H. Gaebler (New York: Academic Press, 1956), p. 119.
H. Goodgal and R. M. Herriott, in The Chemical Basis of
Heredity, ed. W. D. McElroy and
· Glass (Baltimore: Johns Hopkins Press, 1957), p. 336. 3 S.
Zamenhof, in The Chemical Basis of Heredity, ed. W. D.
McElroy and B. Glass (Baltimore: Johns Hopkins Press, 1957),
p. 351. 4A. D. Hershey and M. Chase, J. Gen. Physiol., 36, 39,
1952. 5 A. M. D. Delbruck Hershey, and Virology, G. S. Stent,
1, 108, in 1955; The Chemical 4, 237, 1957. Basis of Heredity,
ed. W. D. McElroy and
· Glass (Baltimore: Johns Hopkins Press, 1957), p. 699. 7C.
Levinthal, these PROCEEDINGS, 42, 394, 1956.
· 2 S.
J. H. Taylor, P. S. Woods, and W. L. Huges, these
PROCEEDINGS, 43, 122, 1957.

26. 9 R. B. Painter, F. Forro,
10 M. S. Meselson, F. W.
Jr., and W. L. Hughes, Nature, 181, 328, 1958.
Stahl, and J. Vinograd, these PROCEEDINGS, 43, 581, 1957.
11 The buoyant density of a molecule is the density of the
solution at the position in the centri fuge cell where the sum of
the forces acting on the molecule is zero. 12 Our attention has
been called by Professor H. K. Schachman to a source of error
in apparent molecular weights determined by density-gradient
centrifugation which was not discussed by In evaluating the
dependence of the free energy of the DNA Meselson, Stahl, and
Vinograd. component upon the concentration of CsCl, the effect
of solvation was neglected. It can be shown that is bound
solvation preferentially. may introduce A method an error for
into estimating the apparent the error molecular due to weight
such selective if either CsCl solvation or water will be
presented elsewhere. 13 In addition to NH4Cl, this medium
consists of 0.049 M Na2HPO4, 0.022 M KH2PO4, 0.05 M NaCl,
0.01 M glucose, 10-3 M MgSO4, and 3 X 10-6 M FeCl3. 14
This result also shows that the generation time is very nearly
the same for all DNA mole This raises the questions of whether
in any one nucleus all DNA mole cules in the population. cules
are controlled by the same clock and, if so, whether this clock
regulates nuclear and cellular division as well.
15 F. H. C. Crick and J. D. Watson, Proc. Roy. Soc. London, A,
223, 80, 1954.
16 R. Langridge, W. E. Seeds, H. R. Wilson, C. W. Hooper, M.
H. F. Wilkins, and L. D. Hamil
ton, J. Biophys. and Biochem. Cytol., 3, 767, 1957.
ed. 18 W. J. D. D. Watson McElroy and and F. B. H. Glass C.
Crick, (Baltimore: Nature, 171, Johns 964, Hopkins 1953. Press,
1957), p. 532. 19 C. E. Hall and M. Litt, J. Biophys. and
Biochem. Cytol., 4, 1, 1958.
17 For reviews see D. 0. Jordan, in The Nucleic Acids, ed. E.
Chargaff and J. D. Davidson (New York: Academic Press,
1955), 1, 447; and F. H. C. Crick, in The Chemical Basis of

27. Heredity,
20 21 R. P. D. Thomas, Lawley, Biochim. Biochim. et Biophys.
et Biophys. Acta, Acta, 14, 21, 231, 481, 1954. 1956. 22 23 S.
Kindly A. Rice supplied and P. by Doty, Dr. J. Michael Am.
Chem. Litt. Soc., The 79, preparation 3937, 1957. of this DNA
is described by Hall
and 24 M. Litt Delbruck, (J. Biophys. these and
PROCEEDINGS, Biochem. Cytol., 40, 783, 4, 1, 1955. 1958).