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![Page 1: It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.](https://reader036.fdocuments.net/reader036/viewer/2022062417/55204d6149795902118b5c22/html5/thumbnails/1.jpg)
„It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanismfor the genetic material.“
„Es ist uns nicht entgangen, dass die spezifische Paarung, die wir vorgeschlagen haben, direkt auf einen möglichen Mechanismus derVervielfältigung des genetischen Materials hinweist.“(James D. Watson und Francis H. C. Crick (1953)
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Wie ist das Zitat von Watson und Crick gemeint? Entwickelt eineHypothese wie die DNA verdoppelt werden kann.
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Versuchsbeschreibung
Meselson und Stahl überprüften 1958 die drei Hypothesen bezüglich der Reduplikation der DNA experimentell
Escherichia-coli-Bakterien (E.-coli) wurden über 14 Generationen in zwei Nährlösungen gezüchtet
Nach drei Tagen wird die DNA aus den Bakterien extrahiert und einer Dichtegradientenzentrifugation
unterzogen.
In einer der Nährlösungen enthält das Ammoniumchlorid (NH4Cl) ein „schweres“ Stickstoffisotop (relative Atommasse 15; 15N).
In der zweiten Nährlösungen enthält das Ammoniumchlorid (NH4Cl) ein „leichteres“ Stickstoffisotop (relative Atommasse 14; 14N).
1
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Dichtegradientenzentrifugation Die Dichtegradientenzentrifugation gehört zu den physikalischen Trennverfahren.
Verschiedene gelöste Makromoleküle werden in einer Ultrazentrifuge anhand ihrer Bewegungsgeschwindigkeit (s.a. Sedimentationsgeschwindigkeit) unter dem Einfluss starker Zentrifugalkräfte sortiert.
Die zu untersuchende Probe wird auf die Oberfläche des Zentrifugenröhrchens gegeben.
Während der mehrstündigen Trennung sedimentieren die Moleküle mit unterschiedlicher Geschwindigkeit im
Lösungsmittel, und zwar solange die Dichte der Probe größer ist als die Dichte des Lösungsmittels und um so
schneller, je größer der Dichteunterschied ist.
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Dichtegradientenzentrifugation Bricht man die Zentrifugation zu einem geeigneten Zeitpunkt ab, erhält man unterschiedliche Banden der Bestandteile der Probe.
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E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
Versuchsdurchführung
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NN
N
NN
N
N N
N
N
N
N
N
NN
Einbau des
„schweren“
Stickstoffes
in die Nucleotide
(Base, Zucker, Phosphat)
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Probenentnahme nach 3 Tagen & DNA-Extraktion
48h Dichtegradientenzentrifugation
Versuchsdurchführung / Phase I
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
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48h Dichtegradientenzentrifugation
? ?
Versuchsdurchführung / Phase I
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
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48h Dichtegradientenzentrifugation
Versuchsdurchführung / Phase I
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
„leichte“ DNA
„schwere“ DNA
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Nach 20 Minuten Dichtegradientenzentrifugation
Überführung in 14NH4Cl Medium („leichten“ Stickstoff)
Versuchsdurchführung / Phase II
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
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Nach 20 Minuten Dichtegradientenzentrifugation
?
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
Versuchsdurchführung / Phase II
4
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Nach 20 Minuten Dichtegradientenzentrifugation
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
Versuchsdurchführung / Phase II
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Transfer auf die molekulare Ebene
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Nach 20 Minuten Dichtegradientenzentrifugation
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
Transfer auf die molekulare Ebene
5
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Transfer auf die molekulare Ebene
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Transfer auf die molekulare Ebene
Generation II
Generation I
Phase I
Phase II
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Generation II
Transfer auf die molekulare Ebene
Generation I
Phase I
Phase II
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Transfer auf die molekulare Ebene
Phase I
Phase II
Generation II
Generation I
6
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Nach 40 Minuten Dichtegradientenzentrifugation
E. coli – Bakterien in 15NH4Cl Medium
E. coli – Bakterien in 14NH4Cl Medium
Transfer auf die molekulare Ebene
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Generation I
Generation II
Generation III
… nach 20 Minuten
… nach 40 Minuten
Phase I
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Transfer auf die molekulare Ebene
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