Manipulating genomes

• What is one application of gene sequencing?


  Genome-wide comparisons between individuals
• How has gene sequencing contributed to comparisons between species?

  It allows for genome-wide comparisons
• What can gene sequencing predict about polypeptides?

  Sequences of amino acids in polypeptides
• What knowledge should you demonstrate about gene sequencing applications?

  • Genome-wide comparisons between individuals
  • Genome-wide comparisons between species
  • Prediction of amino acid sequences in polypeptides
• What is DNA sequencing used for?

  To determine the sequence of nucleotides
• What is Sanger sequencing?

  A method to determine DNA nucleotide sequences
• What components are mixed in Sanger sequencing tubes?

  Primers, DNA polymerase, and free nucleotides
• What happens during the PCR process in Sanger sequencing?

  DNA polymerase synthesizes complementary DNA strands
• What is the result of incorporating chain-terminating nucleotides?

  It stops DNA synthesis at specific bases
• What is produced after many cycles of PCR in Sanger sequencing?

  Thousands of DNA fragments of varying lengths
• What is pyrosequencing?

  A method for sequencing DNA using light detection
• What is fixed to the flow cell in pyrosequencing?

  Single-stranded DNA (ssDNA)
• What happens when activated nucleotides are added in pyrosequencing?

  They are incorporated if complementary to the template
• What is released when a nucleotide is incorporated in pyrosequencing?

  Diphosphate (PPi)
• What does luciferase do in pyrosequencing?

  Converts ATP to oxyluciferin, generating light
• How are unincorporated nucleotides handled in pyrosequencing?

  They are degraded by apyrase
• What is genetic fingerprinting?

  A method to produce DNA band patterns
• What are variable number tandem repeats (VNTRs)?

  Short, repeating sequences in non-coding DNA
• Why is the probability of two individuals having the same VNTRs low?

  VNTRs vary in length and number of repeats
• What are the steps in DNA fingerprinting?

  1. Extraction of DNA & amplification using PCR
  2. DNA digestion with restriction endonucleases
  3. Separation of DNA fragments by gel electrophoresis
  4. Hybridisation with radioactive or fluorescent probes
  5. Development and visualization of banding pattern
• How can DNA profiles be used in forensic science?

  By comparing profiles of suspects and crime scenes
• How can DNA profiles assist in medical diagnosis?

  By identifying individuals at risk of diseases
• What is PCR used for?

  To amplify DNA by artificial replication
• What components are required for PCR?

  DNA sample, nucleotides, Taq polymerase, primers
• What is electrophoresis used for?

  To separate DNA fragments by size
• How does DNA move during electrophoresis?

  It moves towards the anode due to charge
• What is the role of DNA probes in electrophoresis?

  To visualize specific DNA sequences
• What is genetic engineering?

  Altering DNA through recombinant DNA technology
• What are transgenic organisms?

  Organisms expressing genes from other species
• How are DNA fragments created for genetic engineering?

  Using restriction endonucleases and reverse transcriptase
• What is the purpose of a vector in genetic engineering?

  To transfer isolated genes into host cells
• What is transformation in genetic engineering?

  Introducing recombinant plasmids into bacterial cells
• What is gene therapy?

  Treating genetic diseases by inserting functional alleles
• What is somatic therapy in gene therapy?

  Altering alleles in body cells only
• What are ethical concerns of germ-line therapy?

  Potential for designer babies and gene impact