Genomics

Question 1 (10 marks)

(a) (4 marks)

• Higher dNTP:ddNTP ratio (1000:1) means ddATP is incorporated less frequently → chains terminate less often early → produces predominantly longer fragments (1)
• Fewer short fragments generated (1)
• Problem: gaps in the "ladder" — some positions near the start won't be represented, so early bases can't be read reliably (1)
• Reading accuracy at short lengths / short-read resolution is lost, making the 100 bp read incomplete at the beginning (1)

(b) (3 marks)

• Chain elongation requires formation of a phosphodiester bond between the 3′-OH of the last nucleotide and the 5′-phosphate of the incoming nucleotide (1)
• ddNTP lacks the 3′-OH group (1)
• Therefore no bond can form with the next nucleotide → polymerisation terminates at that point (1)

(c) (3 marks) — any two, 1.5 each (round to 3):

• Sanger reads only ~500–1000 bp per reaction → 3 billion bp needs enormous numbers of separate reactions = slow/labour-intensive (1.5)
• Cost per base far higher; NGS is massively parallel (millions of reads simultaneously) making it faster and cheaper (1.5)
• (Accept: throughput of NGS enables whole genome in days, Sanger would take years)

Question 2 (12 marks)

(a) (3 marks)

• Exome = 1.5% of 3200 Mb (1)
• $0.015 \times 3200 = 48$ Mb (2 for correct calculation & answer)
• Answer: 48 Mb

(b) (5 marks) Advantages (2 × 2 = 4, capped appropriately):

• WES targets only ~1.5% (48 Mb) → far less data, lower cost, faster analysis (2)
• Exome contains ~85% of known disease-causing mutations → high diagnostic yield for coding-region disorders (2) Disadvantage (1):
• WES misses ~15% of disease variants located in non-coding/regulatory/intronic regions (1)

(c) (4 marks)

• WES only captures exons (coding regions); a deep intronic variant lies outside the captured/enriched regions so it is not sequenced (2)
• ENCODE showed a large fraction of non-coding DNA is biochemically functional — regulatory elements, enhancers, promoters, transcription-factor binding sites (1)
• Therefore intronic/regulatory variants can affect gene expression and cause disease, justifying WGS in such cases (1)

Question 3 (10 marks)

(a) (2 marks)

• Expected false positives = $0.05 \times 1{,}000{,}000$ (1)
• $= 50{,}000$ SNPs by chance (1)

(b) (4 marks)

• Testing 1,000,000 SNPs = 1,000,000 statistical tests (multiple testing problem) (1)
• At $p<0.05$, 50,000 appear significant purely by chance → unusable (1)
• A stricter threshold corrects for multiple comparisons (Bonferroni-type: $0.05/10^6 = 5\times10^{-8}$) (1)
• This controls the false-positive rate so that "significant" hits are very likely true associations (1)

(c) (4 marks)

• The concept is linkage disequilibrium (1)
• The non-causal SNP is physically close on the chromosome to the true causal variant and is inherited together with it (non-random association) (2)
• So the marker SNP "tags" the causal variant and can be used to locate the disease-associated region even though it isn't itself functional (1)

Question 4 (10 marks)

(a) (4 marks)

• Patient A (normal alleles): normal CYP2C9 activity → normal/faster metabolism → drug cleared at expected rate, normal blood concentration (2)
• Patient B (reduced-function alleles): reduced enzyme activity → slower metabolism → drug accumulates → higher blood concentration, greater risk of toxicity/side effects at standard dose (2)

(b) (3 marks)

• Pharmacogenomics = study of how an individual's genome affects drug response (1)
• Genotyping Patient B beforehand reveals reduced-function alleles (1)
• Allows dose reduction or alternative drug → avoids toxicity and improves safety/efficacy (1)

(c) (3 marks)

• Traditional medicine gives the same dose to all patients regardless of genotype (1)
• Precision medicine tailors drug/dose to the individual's genetic makeup (1)
• Improves efficacy and reduces adverse reactions — treatment matched to the patient (1)

Question 5 (8 marks)

(a) (3 marks)

• Conserved sequences change little over evolutionary time because mutations there are selected against (2)
• This implies the region has an essential function (e.g. vital protein/regulatory role) that must be preserved (1)

(b) (3 marks) — any two:

• Look for open reading frames (ORFs) — start codon, coding sequence, stop codon (1.5)
• Match to known genes/proteins via homology/database (BLAST) alignment (1.5)
• (Accept: presence of promoters/splice sites; evidence from transcriptome/cDNA/RNA-seq showing the region is transcribed)

(c) (2 marks)

• Genome = complete set of DNA/genes (same in all cells) (⅔)
• Transcriptome = all RNA molecules transcribed (varies by cell/condition) (⅔)
• Proteome = complete set of proteins expressed (varies by cell/condition) (⅔)

[
  {"claim": "Exome size = 1.5% of 3200 Mb = 48 Mb", "code": "result = (0.015*3200 == 48)"},
  {"claim": "Expected chance-significant SNPs = 0.05 * 1,000,000 = 50000", "code": "result = (0.05*1000000 == 50000)"},
  {"claim": "Bonferroni threshold 0.05/1e6 = 5e-8", "code": "result = (Rational(5,100)/1000000 == Rational(5,100000000))"},
  {"claim": "Genome-wide threshold equals 5e-8", "code": "result = (5e-8 == 0.05/1000000)"}
]