DNA & Genomics I

Question 1

Nucleotide Structure

Answer 1

1. Pentose 5-carbon sugar
2. Phosphate groups attached to C5 (thus 5’)
3. Nitrogenous bases
   - Purines: Adenine and guanine
   - Pyrimidines: Cytosine and thymine/uracil
   Nucleotide: [nucleoside] __phosphate

Question 2

Nucleic Acid Drawing

Answer 2

Polymerisation reaction with 3’ OH group forming ester linkage with 5’ phosphate group
- Nucleotide: Correct base, phosphate number and deoxyribose/ribose
- Phosphodiester bond
- Hydrogen Bonding between bases, A=T, C three bonds G
- Antiparallel and directionality 5’ -> 3’

Question 3

Structure + Function of DNA

Answer 3

1. Weak hydrogen bonding allows 2 strands to separate and act as templates via CBP to synthesise 2 identical DNA molecules
   - DNA can be accurately replicated w daughter cells having identical DNA copies
2. Strong covalent phosphodiester bond + Numerous H bonds
   - Stable molecule to be passed on to next gen w/o loss of genetic info
3. dsDNA allows 1 strand to be used as template for repair
   - Backup code present with 2 strands
4. Weak H bonds via CBP allow for faithful transfer of info
   - Coded information can be readily utilised

Question 4

Messenger RNA

Answer 4

• Freely mobile mRNA acts as carrier of genetic code from genes in nucleus to ribosomes in cytoplasm/RER, since its synthesised by gene transcription
• mRNA contains triplet codons that code for an AA via CBP, as A=U, T=A and C=G, thus serving as template for translation
• mRNA has start/stop codons and recognition sites to bind to small ribosomal subunit
• Gene expression can be regulated by varying rate on mRNA synthesis

Question 5

Transfer RNA

Answer 5

• Exist in single stranded form but folds back upon itself and held in shape by hydrogen bonds between CBP to form 3D structure
• 3 specific bases form an anticodon that bind to specific mRNA codon via CBP
• 3’ end w CCA stem is attachment site for specific AA
• Function = link to amino acid activation

Question 6

Ribosomal RNA

Answer 6

• rRNA in small ribosomal subunit binds to mRNA via CBP to translocate it
• Main constituent of P and A sites to enable binding of aminoacyl tRNAs
• rRNA also forms peptidyl transferase, a ribozyme catalysing peptide bond formation

Question 7

Semi-Conservative DNA Replication

Answer 7

1. DNA replication is semi conservative, where both original strands separate and serve as templates to synthesise 2 new hybrid DNA molecules made of 1 original and 1 newly synthesised strand
   Start
2. Replication begins at origin of replication
3. Helicase unzips and separates parental stands by breaking H bonds between CBPs
4. Replication forks form and spread in both directions, creating a replication bubble
5. Single-stranded DNA binding proteins bind to separated DNA strands and prevent them from reannealing, remaining single stranded and serving as template
6. Topoisomerase relieves overwinding strain by breaking, swivelling and rejoining DNA strands

Synthesis
7. Short RNA primer added by primase to provide free 3’ OH end
8. DNA polymerase uses parental strand as template and aligns free dNTPs via CBP —> A and T forms 2 H bonds, G and C forms 3 H bonds
9. DNA polymerase catalyses formation of phosphodiester bonds
10. New DNA strand synthesised in 5’ —> 3’
11. DNA strand also proof-reads to ensure proper base pairing
Leading vs Lagging Strand
12. Leading strand synthesised continuously in 5’ —> 3’ direction
13. Lagging strand synthesised discontinuously in short Okazaki fragments in 5’ to 3’ direction as DNA cannot synthesise 3’ to 5’ as active site is only complementary to free 3’ OH group
End
14. A different DNA polymerase removes RNA primer and DNA ligase forms phosphodiester bond to seal the nick
15. Replication bubbles meet to form 2 separate DNA double helixes as complementary parental and newly synthesised DNA strands rewind.

Question 8

Meselson-Stahl Experiment

Answer 8

Parental strand: All heavy 15N-15N DNA
1st generation: All hybrid 14N-15N DNA
2nd generation: 50% 14N-14N + 50% 14N-15N