The Genomics Toolbox and Clinical Genomics in Aotearoa

This lecture covers the laboratory techniques underpinning modern genetics and genomics, and their clinical applications, with reference to the Aotearoa New Zealand context.

THE GENOMICS TOOLBOX

Polymerase Chain Reaction (PCR): amplification of a specific DNA sequence using a thermocycler: denaturation (94C), annealing (50-65C — primers bind target), extension (72C — Taq polymerase synthesises). Exponential amplification produces millions of copies. Applications: pathogen detection (SARS-CoV-2, TB, STIs), mutation detection, STR profiling.

Sanger sequencing: gold standard for sequencing individual genes (~600-1000 bp per reaction). Incorporates fluorescently labelled dideoxynucleotides (ddNTPs) which terminate chain elongation; capillary electrophoresis separates fragments by size. Used for: confirming NGS variants, targeted single-gene testing, Sanger confirmation of novel variants.

Next-Generation Sequencing (NGS): simultaneous sequencing of millions of DNA fragments. Whole Exome Sequencing (WES): all protein-coding exons (~1-2% of genome). Whole Genome Sequencing (WGS): entire genome — detects non-coding variants and structural variants. Gene panels: targeted sequencing of clinically relevant genes (e.g., hereditary cancer: BRCA1/2, MLH1, MSH2; cardiomyopathy; epilepsy).

Array CGH (Comparative Genomic Hybridisation): detects copy number variants (deletions, duplications) across the genome. Resolution ~50-100 kb — detects submicroscopic microdeletion syndromes invisible to karyotyping. First-line for unexplained developmental delay, intellectual disability, and autism.

Karyotype (G-banding): metaphase chromosomes stained and counted; resolution ~5-10 Mb. First-line for confirming suspected aneuploidy (trisomy 21, Turner, Klinefelter), haematological malignancies (Philadelphia chromosome in CML), and large structural rearrangements.

FISH (Fluorescence in situ hybridisation): fluorescently labelled probes hybridise to specific chromosomal loci. Rapid, targeted detection of microdeletions (22q11.2 DiGeorge syndrome), translocations (BCR-ABL1), and gene amplification (HER2 in breast cancer — guides trastuzumab therapy).

GWAS (Genome-Wide Association Studies): compare allele frequencies at hundreds of thousands of SNPs between cases and controls to identify associations with complex traits. Identified thousands of disease loci. Most hits are in non-coding regions; causal variants often unknown. Polygenic risk scores (PRS) aggregate GWAS signals to estimate individual risk — currently limited by population bias (most GWAS from European cohorts) and modest individual predictive value.

NIPT: cell-free fetal DNA in maternal plasma analysed from ~10 weeks gestation. >99% sensitivity/specificity for trisomy 21, 18, 13. A screening test — positive results require confirmatory invasive testing (amniocentesis, CVS) for definitive karyotype.

AOTEAROA NZ CONTEXT

Genomics Aotearoa supports genomic research and implementation in NZ. Key considerations: (1) Population diversity — Maori and Pacific peoples have distinct genomic profiles; variant databases built from European populations may misclassify variants as VUS in Maori patients, causing inequity. (2) Whakapapa (genealogy/lineage) is central to Maori identity — genomic information has collective whanau implications, not only individual ones, challenging individualistic Western consent models. (3) Maori data sovereignty (Te Mana Raraunga): Maori communities assert collective governance rights over genomic data from Maori participants. (4) Equity in access: disparities in genomic testing access for Maori and Pacific peoples require targeted strategies. (5) Pharmacogenomics variants in NZ populations — CYP2C19 poor metaboliser variants relevant to clopidogrel activation are more prevalent in some Pacific peoples.