A research team used the CRISPR-Cas9 system to knock out a gene encoding a cell surface receptor in human embryonic kidney cells. After treatment, they confirmed that the target gene was no longer expressed. Explain how the CRISPR-Cas9 system is able to locate and disrupt a specific gene within the genome of these cells.

OCR A-Level Biology A (H420) — 6.3 Manipulating genomes · Explain · 4 marks · View as Markdown

Written & reviewed by James Millett — Biology (Imperial College London), PGCE Science (University of Cambridge).

CRISPR-Cas9 is a genome-editing tool derived from a bacterial immune defence mechanism. It consists of a guide RNA (gRNA) component and the Cas9 endonuclease enzyme. Scientists design the gRNA to match a target DNA sequence of interest before introducing the complex into cells.

Model answer (4 marks)

The guide RNA (gRNA) is designed to be complementary to a specific DNA sequence in the target gene.
The gRNA binds to that DNA by base‑pairing, directing the Cas9 enzyme to the correct locus.
Cas9 then cleaves both strands of the DNA, creating a double‑strand break at the target site.
The break is repaired by non‑homologous end joining, an error‑prone process that introduces indels, disrupting the reading frame and abolishing gene expression.

Examiner tips

  • Use the exact terms: guide RNA, complementary base pairing, Cas9 endonuclease, double‑strand break, non‑homologous end joining, indels, reading frame disruption.
  • Show the sequence of events in order – design, binding, cutting, repair – to hit all four marks.

Common mistakes

  • Confusing gRNA with Cas9; describing only the cutting step without repair; using vague terms like ‘mutation’ instead of ‘indels’ or ‘reading‑frame disruption’.

Mark scheme (4 marks)

  1. The guide RNA (gRNA) is complementary in base sequence to the target DNA sequence within the specific gene
  2. The gRNA binds to the target DNA sequence by complementary base pairing, guiding the Cas9 enzyme to the correct locus
  3. Cas9 acts as an endonuclease that cuts both strands of the DNA double helix at the target site, producing a double-strand break
  4. The double-strand break is repaired by non-homologous end joining (NHEJ), an error-prone process that introduces insertions or deletions (indels), disrupting the reading frame and preventing expression of the gene

Key terms in this question

CRISPR-Cas9 · genome

Related

More Manipulating genomes questions

▶ Try answering this question with AI marking (free) →