08 Precision Medicine - Human Genome, Genetic Variation and Methods to Detect Variation

Question 1

How many bp does the human genome have?

Answer 1

~3 billion bp

Question 2

What do regulatory elements in a gene do?

Answer 2

control gene expression

Question 3

Properties of a functional genome

Answer 3

• organized
• usable (i.e. regulated and expressed)
• stable
• copied accurately to the next generation of cells

Question 4

What accounts for most phenotypes that are observed?

Answer 4

Genetic variation

Question 5

Types of genetic variations

Answer 5

• Copy number variation
• Structural variation
• Sequence level variation

Question 6

Example of copy number variation

Answer 6

Down syndrome (trisomy 21)

• characteristic facial features
• physical growth delays
• mild to moderate intellectual disability

Question 7

Types of structural genetic variation

Answer 7

• Deletion
• Duplication
• Inversion
• Translocation

Question 8

Example of structural genetic variation

Answer 8

Facioscapulohumeral Muscular Dystrophy (FSHD)
- adolescent onset of progressive weakness of muscles in the face, shoulders and arms
- deletion of ~30-3000kb
- <11 copies of D4Z4 -> expressed to produce DUX4 (toxic to muscles)

Question 9

Types of sequence level variation

Answer 9

• Single nucleotide polymorphism (SNP)
• Insertion/deletion (indel)

Question 10

Example of SNP disease

Answer 10

Sickle cell disease

Question 11

What are dideoxynucleotides?

Answer 11

Nucleotides that lack both the 2’ and 3’ OH group of ribose

Question 12

How does Sanger sequencing work?

Answer 12

• dNTPs are mixed with colored ddNTPs
• Chain termination (no 3’ OH -> cannot form phospodiester bond)
• Separation by size
• 500-800 bp

Question 13

Examples of next generation sequencing technology

Answer 13

Short reads
- Sequencing by synthesis (Solexa/Illumina)
- Nanoball sequencing (Complete Genomics/BGI)
- pH sensing (Ion Torrent/ThermoFisher)

Long reads
- Single molecule sequencing via dye labels (Pacific Bioscience)
- Single molecule sequencing via nanopore sensing (Oxford Nanopore)

Question 14

Limitation of sequencing by synthesis

Answer 14

Sequencing reads are short

Question 15

Why are adaptors and barcodes added to the DNA fragment library before sequencing?

Answer 15

Adaptors are critical for the attachment to the flow cell and allow for the annealing of primers for bridge amplification and the sequencing reaction. Barcodes allow the pooling of multiple sequencing samples in a single sequencing run.