A-Level Biology May/June 2025 Q5(a): To produce a human protein for treatment of a disease, recombinant DNA technology needs…

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  There are several methods to obtain a gene coding for a human protein.

  One method involves using reverse transcriptase. First, mRNA is extracted from a human cell that is actively synthesising the desired protein (e.g., mRNA for insulin from pancreatic β-cells). The enzyme reverse transcriptase is then used to synthesise a single strand of complementary DNA (cDNA) using the mRNA as a template. Following this, DNA polymerase is used to build the second DNA strand, creating a double-stranded DNA copy of the gene. This method is advantageous as the resulting gene lacks introns.

  Alternatively, the gene can be isolated directly from human genomic DNA. The specific gene is located and cut out from the human chromosome using a specific restriction enzyme.

  A third method is to find the published nucleotide base sequence of the gene from an online database. The gene can then be synthesised chemically from individual nucleotides using a gene machine.

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  How the marks are awarded

  • MP4 — Awarded for stating that mRNA is extracted from a cell that is actively making the specific protein, such as pancreatic cells for insulin.
  • MP5 — Awarded for correctly identifying the use of the enzyme reverse transcriptase to produce cDNA from the mRNA template.
  • MP7 — Awarded for describing the alternative method of cutting out or isolating the gene directly from the source DNA, such as a human chromosome.
  • MP8 — Awarded for specifying that a restriction enzyme is used to cut the gene out of the DNA.

  Common mistakes

  • Confusing the roles of enzymes, for example, stating that DNA polymerase is used to make cDNA or that reverse transcriptase cuts DNA.
  • Failing to specify that mRNA must be extracted from a specialised cell type that is actively expressing the gene, instead vaguely saying 'from a human'.
  • Suggesting the use of restriction enzymes to obtain a gene for expression in bacteria, without acknowledging that the resulting gene will contain introns, making it unsuitable.
  • Omitting the DNA polymerase step after reverse transcription, resulting in an answer that only describes the creation of single-stranded cDNA.

  Examiner tip: For process-based questions, memorise the key components—such as specific enzymes and starting materials—and their precise sequence of action.

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