Genetics translation

DNA replication must be

copied with high fidelity relatively quickly

DNA replication model

Semi-Conservative model

Original nuclotide strands remains

intact despite no longer being in the same cell

Original DNA molecule

is halved during replication

Semi conservative model was discovered by

Messelson and Stahl

Messelson and Stahl uses

Centrifugation and radiolabelling nitrogen

Eukaryotes have

linear chromosomes

Prokaryotes

Circular chromosomes

Plasmids

exist outside of the original genome; can replicate on its own and contains antibiotic resistance genes

Replication begins

origin of replication

Replication direction

5’to 3’

Replication bubble

Area where replication happens

Bidirectional

Both directions replication

Unidirectional

one direction replication

Replication fork

the point where DNA unwinds

Theta replication common in

circular DNA

Theta replication replication forks?

one or two(uni or bidirectional)

Theta replication how many origin of replication?

one

Rolling circle replication observed in

Viruses and plasmids

Rolling circle replication initated by

a single break

Rolling circle direction

unidirectional

Yes or no Rolling circle has replication bubble

No

Yes or no multiple replicates can be created quickly

Yes

in what organism does linear replication occur?

Eukaryotes

How many origins of replication in linear replication

multiple

Linear replication Uni or bidirectional

bidirectional

Does Linear replication have replication bubble?

Yea

How many replication forks does linear replication

2

WHat is required for replication

A template consists of a single strand DNA

Raw materials dNTP to be assembled into the new DNA strand

Enzymes and other proteins that read the template and synthesisze the new strand

Dna polymerases can only add

nucleotides to the 3’

What template strand that is exposed 3’→5"‘ called

Leading strand

What template strand that is exposed 5’ → 3’ strand called

Laggins strand

Leading strand has what type of replication?

Continuous

Laggin strand has what type of replication?

discontinuous

What synthesizes the lagging strand?

Okazaki fragments

Where are okazaki fragments found?

found in theta and linear replication

What joins the okazaki fragments?

DNA ligase

What are the 4 steps of replication for prokaryotes

Initation,Unwinding,Elongation,Termination

what is the single replication origin called

OirC

What is the initiator protein called?

DnaA in ECoil

Dna A binds to

oirC and begins to unwind DNA

Does the binding of protein with the oirC signal to additional proteins to join in?

Yes

Dna Polymerase need

a single stranded DNA template

DNA helicase

breaks H-Bonds seperating strands: Needs head start from dnaA and moves 5 → 3 and moves with replication fork

Single stranded binding proteins

protect unwound DNA from forming secondary structures; prevents DNA from re-associating

Dna gyrase

topoisomerase: reduces torque strain that builds ahead of replication fork; breaks and remakes DNA strands through double stranded breaks

Primerase

Synthesizes short stretches of RNA nucleotides to get DNA replication started

Primer on leading strand

one is required

Primer of lagging strand

needs a lot of primers and forms a helicase and moves along the lagging strand together

DNA polymerase III

Multi protein complex that does most of DNA replication: has 5→3’ polymerase activity and has 3→5’ exonuclease activity

DNA polymerase I

used to remove RNA primers; Can synthesize from 5→3 and has exonuclease from 5→3 and 3→5

DNA polymerase II,IV,V

function in DNA repair

What is needs for polymerase

1.Synthesize sequence complementary for the template strand

2. 5→ 3

3. Uses DNTP

4. Needs primers

5. Catalyze the formation of phosphodiester bonds between new nucleotide and growing strand

Dna Ligase in elongation

joins the DNa polymerase I synthesized strand to the DNA polymerase III synthesized strand

Termination occurs

when two replication forks meet

Termination can also occur

at specific sequences where TUS prevents helicase from moving fowards and stalls the replication fork

Initiator protein

binds to the origin and seperates strands of DNA to initiate replication

DNA helicase

Unwinds DNA at replication fork

SSBP

attaches to the single-stranded DNA and prevents secondary structure from forming

DNA gyrase table def

moves ahead of the replication fork making and resealing breaks in the DNA to release torque

DNA primase

attaches a RNA primer to provide a 3’OH group for the attachment of DNA nucleotides

DNA polymerase III

elongates a new nucleotide strand from the 3’OH group provided by the primer

DNA polymerase I

Removes RNA primers and replaces

DNA ligase

joins okazaki fragments by sealing nicks

Eukaryotes Origins of replication calles

autonomously replicating sequences

Initations has two parts in order

ensure that all DNA Is replicated at the same time

First step of initation in eukaryotes

replication licensing factors

Second step of initation in eukaryotes

Replications occurs at licensed sites

how many polymerases does eukaryotes have

multiple DNA polymerases

Eukaryotes must reassemble

nucleosomes

Three polymerases in eukaryotes

alpha,delta, and epsilon

PCR allows

thousands of copies of DNA produced very quickly and outside of the cell

PCR allows

specific gene to be ampified

Where does the PCR occur

Thermalcycler

three steps of PCR

Denaturation,Annealing,Extension

Denaturation

occurs using hear(95C)

Annealing

60 C. Temp is lowered to allow primers to anneal

Extension

created a new strand of DNA starting on the primer with taq polymerase

the sequence of your primers determines

location of the gene that is going to be amplified, specificity of PCR, efficiency and annealing temperature

Annealing temp controls

stringency

high stringency

only one product can be made in PCR

High stringecny requires

primer to be an exact match to the DNA

High stringency Temp

High annealing temperature

Low stringency

multiple products can be made

Low stringency primer

product is similar to the primer

Low stringency temp

Low

Low stringency use?

want to amplify a family of related genes in a genome

High stringency length

long primer (20-25) to ensure that primer is an exact match

Low stringency length

5-15. Short primer. higher probability of repeating in genome. Multiple products