DNA Organization and Replication Genetics Unit I-Part B DNA Organization and Replication

An Introduction “A genetic material must carry out two jobs: duplicate itself and control the development of the rest of the cell in a specific way.” -Francis Crick "It has not escaped our notice that the specific (base) pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.“ -Watson and Crick April 25, 1953

the fun’s just getting underway Eukaryotic DNA Organization DNA is very organized. The amount of DNA packed into a nucleus would equal 5 miles of string packed into a film cannister The double helix is wrapped around protein particles called nucleosomes. These nucleosomes are made up of 8 proteins called histones. the DNA then resembles “beads on a string” This allows the DNA to be very tightly packed and organized These nucleosomes must be unwound in order for the DNA to replicate

Nucleosome diagram

Replication of the Genetic Material Replication is defined as the process by which the genetic material is reproduced, leaving 2 exact copies to be passed on to the daughter cells during mitosis. For many years scientists debated how replication took place. Three models were generally discussed:

Models for DNA replication 1) Semiconservative model: Daughter DNA molecules contain one parental strand and one newly-replicated strand 2) Conservative model: Parent strands transfer information to an intermediate, then the intermediate gets copied. The parent helix is conserved, the daughter helix is completely new 3) Dispersive model: Parent helix is broken into fragments, dispersed, copied then assembled into two new helices. New and old DNA are completely dispersed

MODELS OF DNA REPLICATION Semi-conservative replication Conservative replication Dispersive replication

Which model is correct? Proven by Meselson-Stahl Experiment Materials used -Ultracentrifuge, bacteria, two types of radioactive nitrogen, 15N (heavy), 14N (light), Cultured bacteria -Method used: Grew bacteria in medium containing heavy nitrogen, until all DNA contained this isotope Next, grew the bacteria in light nitrogen for 1 cell division, the DNA was then isolated and centrifuged The DNA bands in the centrifuge were examined, results analyzed

Which model is correct? This experiment was continued over further generations, the results showed replication took place in a semi-conservative manner.

Features of DNA Replication DNA replication is semi-conservative Each strand of template DNA is being copied. DNA replication is bidirectional Bidirectional replication involves two replication forks, which move in opposite directions DNA replication is semi-discontinuous The leading strand copies continuously The lagging strand copies in segments (Okazaki fragments) which must be joined DNA replication involves many enzymes and factors

DNA Replication is bi-directional

Discontinuous synthesis DNA replication is semi-discontinuous Discontinuous synthesis

Notes to diagram Okazaki showed that the 2nd strand is replicated discontinuously, in short pieces. 1st, RNA primers are synthesized, then DNA is added. The RNA primers are then degraded + DNA is filled in. The pieces are called Okazaki fragments.

DNA replication involves many enzymes and factors The method of DNA replication Many enzymes + factors are involved in the process. The main class of enzymes are the DNA polymerases (DNA pol’s)

The DNA Polymerase Family A total of 5 different DNAPs have been reported: DNAP I: functions in repair and replication DNAP II: functions in DNA repair (proven in 1999) DNAP III: principal DNA replication enzyme DNAP IV: functions in DNA repair (discovered in 1999) DNAP V: functions in DNA repair (discovered in 1999)

DNA replication involves many enzymes and factors Other Enzymes and Factors Needed 1.) helicase – enzyme which unwinds the DNA double helix 2.) RNA primase – enzyme which makes the RNA primers 3.) DNA ligase – enzyme which binds Okazaki fragments together 4.) single-stranded binding proteins (SSB’s) – keep single-stranded DNA stable when it’s unwound 5.) gyrase – prevents DNA from tangling as it unwinds (aka the “Conditioner” enzyme