(1 - 2) - Leading Strand With Helicase - Lagging Strand Opposite Heliase

Topic 2: Molecular Biology - 2.
DNA Replication is the process in which present DNA is copied such that precisely the same DNA
can be used to allow cells to be made by mitosis for growth and Repair, for example. DNA is copied
using the “semi-conservative” method and functions through many steps by enzymes. DNA
replication is only possible due to complementary pairing and is, therefore, explained accordingly:
DNA Replication: Step by Step
- The Double Helix is unzipped by the Enzyme Helicase

- Hydrogen Bonds are Broken
- Helicase works step by Step - Like slowly Unzipping Zipper
- The Nitrogenous bases are Unpaired
- In the Environment, there are Free Nucleotides
- Are available to Form Complementary Pairs
- DNA Polymerase -> Matches Free Nucleotides to Bases
- 2 New Identical DNA Strands are Formed; Replication
- Splitting Point = Replication Fork

- Like Road Fork (1 -> 2)
- The direction of DNA Synthesis
- Leading Strand = With Helicase
- Lagging Strand = Opposite Heliase
- Primers = Place Holders
- Only for Lagging Strand
- New Strand = ½ Parent Strand
- Semi-Conservative
- From 1 Parent DNA -> 2 Identical DNA
Terminology
Helicase Enzyme that Splits DNA into its 2 Independent Strands - Needs ATP
DNA Polymerase Enzyme that catalyzes Free Nucleotides with Split Strands
Replication Fork The point in which DNA is Split into Two like a Road Fork
Leading Strand The Strand which is Continuously Replicated - Same Direction as Helicase
Lagging Strand Strand which is not Continuously Replicated - Opposite Direction as Helicase
DNA Polymerase creates Complementary Strands: More Detailed
DNA Polymerase moves in a 5 Prime to 3 Prime Direction
DNA Polymerase Catlyses the covalent popphodiester bonds between sugars and Phosphates
DNA Polymer proofreads the Complementary Base Pairing; Mistakes are very Infrequent
Free nucleotides are deoxynucleoside triphosphates; Extra phosphate = Energy for covalent bonds
Meselson and Stahl: Semi-Conservative Replication
- Mesolson and Stahl grew E-Coli in Heavy Nitrogen (15N)

- Since DNA constraints Nitrogen, 15N became incorporated into DNA
- Bacteria were Transferred to light nitrogen 14N
- All new DNA from this point would contain light nitrogen
- After Each Generation samples where Taken
- Samples were analyzed using Density gradient Centrifugation
- Separates DNA based on Density
- More Dense = Lower in Tube
- Less Dense = Higher in Tube
- Compared 15N and 14N with DNA Generations
- Found that new Generations where a Intermediate between 15N and 14N
- Indicated a hybrid between both Light and Heavy Nitrogen
- Proof that DNA Replication is Semi-Conservative
Polymerase Chain Reaction: PCR
Polymerase Chain Reaction essential is the synthetic comparative to DNA replication. PCR is
Typically used to copy a specific segment of DNA – not a whole genome. PCR amplifies small
samples of DNA To use them for DNA profiling, recombination, species identification, or other
research when not a large sample is present or available.
Process Explanation:
The process requires; Thermal cycler, primers, free DNA nucleotides, and DNA polymerase.
PCR occurs in a thermal cycler and involves a repeat procedure of 3 steps:

1. Denaturation: The DNA sample is heated to separate it into two strands (1)
2. Annealing: DNA primers attach to opposite ends of the target sequence (2)
3. Elongation: A heat-tolerant DNA polymerase (Taq) copies the strands (3)
a. Free Nucleotides are Joined through the Energy of the Third Phosphate
(1) = 95°C | (2) = 65°C | (3) = 72°C
One cycle of PCR yields two identical copies of the DNA sequence
A standard reaction of 30 cycles would yield around 1 Billion copies of DNA
Why do we use Taq Polymerase?
Because of the heating and cooling in PCR, Polymerase from the bacteria Thermus Aquaticus lives in
hot springs and hydrothermal vents. This works because DNA code is universal - The sequence of
GTCA is found within all organisms. At 95 degrees celsius, human polymerase won't function
because this would cause the enzyme to denature due to the excessive temperature; Hence another
Enzyme is used
Note: This is Evidence that the DNA code is Universal !!!

Transcription and Translation: (This is independent of Replication)
- (1) Transcription = To make an mRNA copy of a specific gene (Gene codes for 1 polypeptide)
- (2) Translation = To use this mRNA copy for a specific polypeptide
- protein Synthesis in Ribosomes
Eukaryotic vs. Prokaryotic: Differences in Process
Structure: Relevance:
- Single Celled Organism

- Transcription = Cytoplasm
Prokaryotes - No Nucleus
- Translation = Cytoplasm
- Small Ribosomes
- Multicellular Organism
- Transcription = Nucleus
Eukaryotes - Large Ribosomes
- Translation = Cytoplasm
- Nucleus
Transcription: Making mRNA Copy of a Specific gene
RNA polymerase separates the DNA strands and synthesizes a complementary RNA copy
This Occurs on the Template Side (Antisense) - The Cell knows which side this corresponds to
RNA Polymerase Covalently bonds ribonucleoside triphosphates - A = U , G = C
Once the RNA sequence has been synthesized = RNA polymerase will detach from the DNA; RNA
detaches from the DNA; the double helix of the DNA reforms -> Hydrogen Bonds Restored
The sequence of DNA that is transcribed into RNA is called a gene

Translation: Utilizing mRNA Strand to synthesize Polypeptides
mRNA strand travels outside of the Nucleus into Cytoplasm -> Translation process can Start
The mRNA will locate a ribosome and align with it -> Ribosome Synthesized in 1 Direction
A specific tRNA molecule nowfloats in; The anticodon of the molecule most fit the mRNA
Ribosome alignes the tRNA and mRNA according to Complementary Base Pairing
tRNA is detached from the Amino Acid -> Amino Acid is condensed in Correct Sequence
A peptide bond is formed between the two amino acids (carried by the tRNAs)
1. mRNA acts as a Blueprint for Code
2. tRNA molecules contain anticodons

which are complementary to the
codons on the mRNA.
3. RNA molecules bind to a specific

amino acid that corresponds to the
anticodon
4. tRNAs with anticodons

complementary to the codons bind
(the bases are linked by the formation
of hydrogen bonds)
Definition and Explanation Table: Additional Information
Structure of tRNA
Anticodon = Opposite Complementary of Codon

↳ Example: Codon = UGAU ∴ Anticodon = ACUA
A Codon is a 3 Letter combination of Nucleotide Bases

↳Universal Language of DNA/RNA
Codon
Start Codon = AUG (Methionine)

End Codon = UAG; UAA; and UGA
Structure Eukaryote
Structure Prokaryote
Sequence Decoding: Skills

(1 - 2) - Leading Strand With Helicase - Lagging Strand Opposite Heliase

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Topic 2: Molecular Biology - 2.

DNA Replication: Step by Step

- The Double Helix is unzipped by the Enzyme Helicase

- Splitting Point = Replication Fork

DNA Polymerase moves in a 5 Prime to 3 Prime Direction

Meselson and Stahl: Semi-Conservative Replication

- Mesolson and Stahl grew E-Coli in Heavy Nitrogen (15N)

PCR occurs in a thermal cycler and involves a repeat procedure of 3 steps:

(1) = 95°C | (2) = 65°C | (3) = 72°C

A standard reaction of 30 cycles would yield around 1 Billion copies of DNA

Why do we use Taq Polymerase?

Note: This is Evidence that the DNA code is Universal !!!

Eukaryotic vs. Prokaryotic: Differences in Process

- Single Celled Organism

Transcription: Making mRNA Copy of a Specific gene

RNA Polymerase Covalently bonds ribonucleoside triphosphates - A = U , G = C

The sequence of DNA that is transcribed into RNA is called a gene

1. mRNA acts as a Blueprint for Code

2. tRNA molecules contain anticodons

3. RNA molecules bind to a specific

4. tRNAs with anticodons

Anticodon = Opposite Complementary of Codon

A Codon is a 3 Letter combination of Nucleotide Bases

Start Codon = AUG (Methionine)

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