DNA – Deoxyribose nucleic acid (double stranded/longer)
Phosphodiester bond joins nucleotides together (sugar phosphate backbone)
Hydrogen bonds between the base pairs form the double strand
DNA provides code for an organism’s proteins.
Sequence of base pairs (unique to each individual) are the code that determines which proteins are manufactures.
3 bases code for one amino acid. The amino acids join to form the polypeptide chain.
Degenerate: The same amino acid can be coded for by more than one base triplet
Universal: A base triplet will code for the same amino acid in all species
Non-overlapping: No bases are shared between triplets
Triplet code: Each amino acid is coded for by three bases
RNA – Ribonucleic acid (single stranded/shorter)
mRNA : a single stranded polynucleotide chain that if formed in the nucleus using a section on the coding strand of the DNA as a template.
Each set of three bases on the mRNA are called a codon.
tRNA : a single stranded polynucleotide chain that is folded into a clover shaped molecule.
Base pairing occurs in certain parts of the molecule which hold the shape if the molecule together. Each molecule has a binding site at one
end with base sequence ACC, where the amino acid attaches itself.
Each also has a specific sequence of three bases at the opposite end of the molecule called an anticodon.
QU. What is the difference between a polypeptide and a protein?
Answer: A polypeptide is a sequence of amino acids (primary structure), and a protein is the final functioning molecule with a specific
shape.
Transcription:
-DNA double helix unwinds at the gene caused by DNA helicase breaking H bonds.
-One strand acts as a template
-Free RNA nucleotides line up opposite their complementary base
-RNA polymerase joins nucleotides together (condensation reactions)
-mRNA leaves nucleus via nuclear pore
mRNA Modification:
Noncoding introns are removed. Exons are joined together. This is why mRNA is shorter than DNA.
(Intronnoncoding protein) (Exon coding protein)
Translation:
-mRNA attaches to a ribosome which holds two codons
-First codon (in the P site) attracts the tRNA with the complementary anticodon (moves into P site)
-Second anticodon (in the A site) also attracts the tRNA with the complementary anticodon.
-The tRNA carries specific amino acid to the ribosome
-A peptide bond forms between the two amino acids
-The ribosome shifts over by one codon and frees up the A site for the next tRNA
Gene Mutations: A change in the base sequence of a gene.
There are four types of gene mutation
- Substitution
- Deletion
- Addition/Insertion (extra base is added to base sequence)
- Inversion (bases within triplet reversed)
Substitution: (E.g., AGT AGA)
Only affect one triplet , so only one amino acid may be changed. In some cases, the change of just one amino acid does not affect the
overall structure and function of the protein.
Deletion: (E.g., AGT ATT CGC AGT TTC GC… )
Far greater affect than substitution, cause major changes in amino acid sequence.
Addition of loss of a base causes a shift in the whole sequence of bases and all triplet codes are changes after that point. This is a frame
shift.
Causes a different primary structure different tertiary structure different shape and no function.
What causes mutations?
Occur randomly and spontaneously, but rate may increase due to carcinogens/radiation
Body cells with multiple mutations can become cancerous. Normal body cell function ceases. Cell divides uncontrollably causing tumour.
, Transgenics
Recombinant DNA technology involves combination of DNA from one organism with DNA from another organism .This means inserting a
gene from one species into another.
Recombinant DNA Technology:
- Locate the human gene using a DNA probe (short, single stranded length of DNA with complementary base sequence to the
gene)
- Isolate the human gene: Cut using restriction endonuclease (cuts from the chromosome at a specific base sequence to create
sticky ends) or make using reverse transcriptase (using mRNA as a template to create DNA, then made double stranded by DNA
polymerase. DNA produced is called cDNA. It is shorter than corresponding gene as introns have been removed after
transcription)
- Prepare the plasmid (vector) by cutting it open, using restriction endonuclease. (Essential because the sticky ends of the cut
plasmid will be comp. to the sticky ends of the cut human gene, which allows the two pieces of DNA to fit together.)
- Insert the human gene into the plasmid. Sticky ends allow complementary base pairing (H bonds). DNA ligase joins DNA
(ligation/splicing). (Ligase enzyme catalyses formation of phosphodiester bond.)
- Place bacteria in cold calcium chloride (increase membrane permeability) and a sudden temperature raise (heat shock) causes
some bacteria to take up plasmid. Bacteria that take up bacteria are called transformed.
- Use marker genes to identify transformed bacteria e.g., Antibiotic resistance genes/Fluorescent marker genes.
- Allow transformed bacteria to reproduce under optimum conditions
- Bacteria will make human protein e.g., insulin
Advantages of making human proteins using bacteria:
- If sourced from other animals, will not be as effective as human sourced; could cause allergies/ viral infections
- Ethical reasons : vegetarians/religions do not use certain animal products
- Bacteria allows for mass production
Definitions:
Sticky ends: Single stranded complementary sequences at the end cut DNA
cDNA: DNA made from mRNA
Vector: Carries human gene into the host cell (E.g., plasmid and viruses)
Recombinant: has DNA from more than one species)
PCR - Polymerase Chain Reaction --> producing multiple copies of a small sample of DNA (amplify the sample DNA)
The Raw Materials for PCR:
-Original DNA sample
-DNA polymerase
-DNA nucleotides (A,T,G,C)
-Primers (short pieces of single stranded DNA with complementary base sequence to the end of the DNA strand)
Process:
- Heat DNA sample to 95 degrees to break H bonds between bases of the two strands of DNA. DNA is now single stranded.
- Primers, DNA polymerase and free DNA nucleotides are added to single stranded DNA.
- Mixture is cooled to 40 degrees to allow binding.
- Mixture is heated o 70 degrees. DNA polymerase joins nucleotides attached free nucleotides together with template DNA strand, to
create new DNA strand.
- Recycle: Each cycle doubles the amount of DNA. E.g., How many DNA molecules after the 6th cycle = 64
QU. Why are primers needed?
Act as a starting point for the formation of the new DNA strand. Allow DNA polymerase to bind on. Prevent the two DNA strands from
reforming the double helix.)
Differences between PCR and DNA replication:
DNA replication uses DNA helicase to separate the strands. PCR uses heat.
DNA replication does not require primers.
*Important there is no contaminating DNA, or it will also be replicate. DNA polymerase enzymes must be thermostable (not denatured at
the high temperatures).
Phosphodiester bond joins nucleotides together (sugar phosphate backbone)
Hydrogen bonds between the base pairs form the double strand
DNA provides code for an organism’s proteins.
Sequence of base pairs (unique to each individual) are the code that determines which proteins are manufactures.
3 bases code for one amino acid. The amino acids join to form the polypeptide chain.
Degenerate: The same amino acid can be coded for by more than one base triplet
Universal: A base triplet will code for the same amino acid in all species
Non-overlapping: No bases are shared between triplets
Triplet code: Each amino acid is coded for by three bases
RNA – Ribonucleic acid (single stranded/shorter)
mRNA : a single stranded polynucleotide chain that if formed in the nucleus using a section on the coding strand of the DNA as a template.
Each set of three bases on the mRNA are called a codon.
tRNA : a single stranded polynucleotide chain that is folded into a clover shaped molecule.
Base pairing occurs in certain parts of the molecule which hold the shape if the molecule together. Each molecule has a binding site at one
end with base sequence ACC, where the amino acid attaches itself.
Each also has a specific sequence of three bases at the opposite end of the molecule called an anticodon.
QU. What is the difference between a polypeptide and a protein?
Answer: A polypeptide is a sequence of amino acids (primary structure), and a protein is the final functioning molecule with a specific
shape.
Transcription:
-DNA double helix unwinds at the gene caused by DNA helicase breaking H bonds.
-One strand acts as a template
-Free RNA nucleotides line up opposite their complementary base
-RNA polymerase joins nucleotides together (condensation reactions)
-mRNA leaves nucleus via nuclear pore
mRNA Modification:
Noncoding introns are removed. Exons are joined together. This is why mRNA is shorter than DNA.
(Intronnoncoding protein) (Exon coding protein)
Translation:
-mRNA attaches to a ribosome which holds two codons
-First codon (in the P site) attracts the tRNA with the complementary anticodon (moves into P site)
-Second anticodon (in the A site) also attracts the tRNA with the complementary anticodon.
-The tRNA carries specific amino acid to the ribosome
-A peptide bond forms between the two amino acids
-The ribosome shifts over by one codon and frees up the A site for the next tRNA
Gene Mutations: A change in the base sequence of a gene.
There are four types of gene mutation
- Substitution
- Deletion
- Addition/Insertion (extra base is added to base sequence)
- Inversion (bases within triplet reversed)
Substitution: (E.g., AGT AGA)
Only affect one triplet , so only one amino acid may be changed. In some cases, the change of just one amino acid does not affect the
overall structure and function of the protein.
Deletion: (E.g., AGT ATT CGC AGT TTC GC… )
Far greater affect than substitution, cause major changes in amino acid sequence.
Addition of loss of a base causes a shift in the whole sequence of bases and all triplet codes are changes after that point. This is a frame
shift.
Causes a different primary structure different tertiary structure different shape and no function.
What causes mutations?
Occur randomly and spontaneously, but rate may increase due to carcinogens/radiation
Body cells with multiple mutations can become cancerous. Normal body cell function ceases. Cell divides uncontrollably causing tumour.
, Transgenics
Recombinant DNA technology involves combination of DNA from one organism with DNA from another organism .This means inserting a
gene from one species into another.
Recombinant DNA Technology:
- Locate the human gene using a DNA probe (short, single stranded length of DNA with complementary base sequence to the
gene)
- Isolate the human gene: Cut using restriction endonuclease (cuts from the chromosome at a specific base sequence to create
sticky ends) or make using reverse transcriptase (using mRNA as a template to create DNA, then made double stranded by DNA
polymerase. DNA produced is called cDNA. It is shorter than corresponding gene as introns have been removed after
transcription)
- Prepare the plasmid (vector) by cutting it open, using restriction endonuclease. (Essential because the sticky ends of the cut
plasmid will be comp. to the sticky ends of the cut human gene, which allows the two pieces of DNA to fit together.)
- Insert the human gene into the plasmid. Sticky ends allow complementary base pairing (H bonds). DNA ligase joins DNA
(ligation/splicing). (Ligase enzyme catalyses formation of phosphodiester bond.)
- Place bacteria in cold calcium chloride (increase membrane permeability) and a sudden temperature raise (heat shock) causes
some bacteria to take up plasmid. Bacteria that take up bacteria are called transformed.
- Use marker genes to identify transformed bacteria e.g., Antibiotic resistance genes/Fluorescent marker genes.
- Allow transformed bacteria to reproduce under optimum conditions
- Bacteria will make human protein e.g., insulin
Advantages of making human proteins using bacteria:
- If sourced from other animals, will not be as effective as human sourced; could cause allergies/ viral infections
- Ethical reasons : vegetarians/religions do not use certain animal products
- Bacteria allows for mass production
Definitions:
Sticky ends: Single stranded complementary sequences at the end cut DNA
cDNA: DNA made from mRNA
Vector: Carries human gene into the host cell (E.g., plasmid and viruses)
Recombinant: has DNA from more than one species)
PCR - Polymerase Chain Reaction --> producing multiple copies of a small sample of DNA (amplify the sample DNA)
The Raw Materials for PCR:
-Original DNA sample
-DNA polymerase
-DNA nucleotides (A,T,G,C)
-Primers (short pieces of single stranded DNA with complementary base sequence to the end of the DNA strand)
Process:
- Heat DNA sample to 95 degrees to break H bonds between bases of the two strands of DNA. DNA is now single stranded.
- Primers, DNA polymerase and free DNA nucleotides are added to single stranded DNA.
- Mixture is cooled to 40 degrees to allow binding.
- Mixture is heated o 70 degrees. DNA polymerase joins nucleotides attached free nucleotides together with template DNA strand, to
create new DNA strand.
- Recycle: Each cycle doubles the amount of DNA. E.g., How many DNA molecules after the 6th cycle = 64
QU. Why are primers needed?
Act as a starting point for the formation of the new DNA strand. Allow DNA polymerase to bind on. Prevent the two DNA strands from
reforming the double helix.)
Differences between PCR and DNA replication:
DNA replication uses DNA helicase to separate the strands. PCR uses heat.
DNA replication does not require primers.
*Important there is no contaminating DNA, or it will also be replicate. DNA polymerase enzymes must be thermostable (not denatured at
the high temperatures).
