3.1 Essential ideas

3.1.5 Genetic modification and biotechnology

Biologists have developed techniques that allow for the artificial manipulation of DNA, cells and organisms.

Polymerase chain reaction

  • The polymerase chain reaction (PCR) is a technology used to amplify small quantities of DNA into large samples that can be further analysed.

Figure 3.1.5a – PCR amplifies small amounts of DNAFigure 3.1.5a – PCR amplifies small amounts of DNA

  • Sequences of DNA are targeted using DNA primers that bind by complementary base pairing to initiate DNA replication.
  • Millions of copies of the targeted DNA sequence can be made within hours.

Gel electrophoresis

  • Gel electrophoresis is used to separate charged molecules (proteins or fragments of DNA) according to their size.
  • The technique uses a gel, immersed in a conducting fluid onto which an electrical field is applied.

Figure 3.1.5b – Gel electrophoresisFigure 3.1.5b – Gel electrophoresis

  • DNA fragments of varying lengths are loaded into wells on the gel at the negative electrode.
  • DNA fragments, being slightly negatively charged, move through the gel towards the positive electrode.
  • Smaller fragments move faster and travel further down the gel than larger fragments.

DNA profiling

  • DNA profiling involves comparing samples of DNA.
  • Every individual has a unique pattern of DNA fragmentation (like a fingerprint), so this technique is useful for forensic investigations, and in cases of unknown paternity.
  • A DNA profile is made by:
    • obtaining a sample of DNA (i.e. from a crime scene)
    • using PCR to amplify the amount of DNA in the sample
    • cutting the DNA into fragments using restriction enzymes that target specific sequences (the resulting fragments will be different lengths)
    • separating the fragments of DNA sample by gel electrophoresis.
    • When samples from different individuals are loaded on the same gel, banding patterns can be compared.

Skill: Analysing DNA profiles

The DNA of two men (F1 and F2) who claim to be the father of a child is compared to the DNA of the mother (M) and the child (C) in the DNA profile below.

Figure 3.1.5c – DNA profiles for determining paternityFigure 3.1.5c – DNA profiles for determining paternity

Source: Griffiths et al, Introduction to Genetic Analysis, 6th edition

  1. Print the profile. Label the negative electrode and the positive electrode.
  2. Circle the longest DNA fragment on the image.
  3. Account for the bands in C by indicating whether the band is common between mother and child, or between father and child.
  4. Determine whether F1 or F2 is the father of C. Explain your choice.
    (Scroll over image for answers.)

Genetic modification

  • Genetic modification, or genetic engineering, involves gene transfer between species.
  • Genes may be transferred to bacterial plasmids, restriction endonucleases and DNA ligase.
  • The bacterial plasmids are then used as vectors to deliver new genes into organisms. Other vectors might be viruses or a recombinant cell.

Figure 3.1.5d – Gene transfer using a plasmid vector in the production of human insulinFigure 3.1.5d – Gene transfer using a plasmid vector in the production of human insulin

  • Restriction endonucleases are enzymes that digest DNA strands at specific sequences, leaving behind single-stranded ‘sticky ends’.

Figure 3.1.5e – Sticky ends bind by complementary base pairingFigure 3.1.5e – Sticky ends bind by complementary base pairing

  • The desired gene is mixed with bacterial plasmids that have been cut with endonucleases. The sticky ends bind by complementary base pairing to the inserted gene. DNA ligase enzyme joins the DNA backbones to produce a recombinant plasmid.
  • The recombinant is cloned in small populations and then on a larger scale to produce the desired gene product.

Figure 3.1.5f – Thermal cyclerFigure 3.1.5f – Thermal cycler
Care must be taken to ensure that samples loaded into a PCR thermal cycler are not contaminated.

Figure 3.1.5g – Industrial fermentation insulinFigure 3.1.5g – Industrial fermentation insulin
Industrial manufacture of insulin at Novo Nordisk, Inc. (Denmark).

TOK

The use of DNA evidence in legal cases is well-established. What criteria are necessary for establishing the reliability of evidence?

Figure 3.1.5h – ElectrophoresisFigure 3.1.5h – Electrophoresis
Agarose gels can be used to perform simple electrophoresis in schools.

Figure 3.1.5i – FluorescentFigure 3.1.5i – Fluorescent
DNA is colourless so fluorescent dyes are used to detect DNA in electrophoresis.

Course links

  • More information on PCR and plasmid gene transfer for the production of human insulin is available in 2.2.7.
  • HL: Details on the specific sequences used in DNA profiling and the role of electrophoresis in genetic sequencing are in 7.2.1.