Genetic Testing and Single Base Variation

Definitions

  • Mutation: any heritable change in the human genome which causes a genetic disorder
  • Polymorphisms: any variation in human genome which has a population frequency >1%
    • Does not cause disease in its own right, but may predispose to common disease
  • Penetrance: the likelihood of having a disease if you have a gene mutation
  • Classical genetic disease (Mendelian disorders): one mutation sufficient to cause disease
    • High penetrance, small environmental contribution
  • Multifactorial disease: multiple polymorphisms cause a risk of disease
    • Penetrance for any one mutation is low

Gene analysis

aCGH

  • 1st line chromosome test
  • Large scale - 3 million BPs
  • Detects missing/duplicated pieces of chromosome
  • Find polymorphisms
  • Does not detect balanced rearrangements

FISH

  • Uses fluorescent probes that bind only to parts of a nucleic acid sequence with a high degree of sequence complementarity
  • Often used for finding specific features in DNA for use in genetic counselling

PCR

  • Can select one small piece of human genome from a patient and amplify it
  • Pieces can be selected to find mutations

Whole exome sequencing

  • Sequences exome - all the exons (vs. whole genome sequencing – introns and exons)

Genetic filtering

  • On average approx. 3 000 000 polymorphisms are detected when sequencing the entire genome in a person
  • To identify the pathogenic variant, the list of variants is filtered to remove those that are unlikely to be disease causing

Types of mutation

Missense mutations

  • Point mutations that cause a change to a single amino acid
  • Usually caused by substitutions
  • Most likely mutation to directly activate an oncogene
    • Other mutations tend to inactivate the gene

Changes to amino acid sequence

  • Mutation may change protein sequence which may alter protein function
  • Mutation may result in a premature stop codon

Insertion/deletion

  • Causes a complete change to the entire amino acid sequence after the mutation site
  • In frame - insertion/deletion of a multiple of 3 bases
  • Out of frame - results in a frame shift

Promotor and splice site sequence changes

  • Stop transcription or cause abnormal splicing

Nomeclature

  • g. (genomic build) – e.g. g.123[G>A];[=]
  • c. (RNA as if DNA) – e.g. c.125[C>G];[=]
  • p. (protein) – e.g. p.Pro172Arg
  • > = sub
  • ins/del e.g. c.76insG
  • c.267+2 = within intron