2 replication big picture

Semiconservative replication

Each daughter duplex has one parental strand and one new strand

Meselson-Stahl setup

Grow E. coli in heavy 15N, switch to light 14N, sample over time, separate DNA by density in a CsCl gradient

Meselson-Stahl result after 1 generation

One hybrid H-L band (rules out conservative, which predicts H-H + L-L)

Meselson-Stahl result after 2 generations

Two bands: H-L and L-L

Origins in bacteria vs eukaryotes

Bacteria (circular): usually one origin. Eukaryotes (linear): many origins

Bidirectional replication

Two replication forks form at the origin and move away in opposite directions

Replication fork

Y-shaped point where parental strands separate and new daughter strands are made

Leading strand

Made continuously in the same direction the fork moves

Lagging strand

Made discontinuously as Okazaki fragments, in the direction opposite fork movement, then joined by ligase

Semidiscontinuous replication

One strand continuous (leading), one discontinuous (lagging)

Okazaki fragment length

1-2 kb in bacteria; 100-200 nt in eukaryotes

Okazaki fragment primer

Short RNA primer of 10-13 nt at the 5' end of each fragment

How Okazaki found the fragments

T4 phage-infected E. coli, brief radioactive labeling, alkaline CsCl gradients

Direction of DNA synthesis

Always 5' to 3' on both leading and lagging strands