Biology • Year 12 • Module 6 • Lesson 15
Cloning, Whole Organism and Gene Cloning
Apply cloning theory to real numbers: the Dolly experiment success rate, the Tasmanian thylacine cloning attempts, and a worked case on gene cloning of human insulin.
1. Dolly the sheep, what the real numbers say about effectiveness
Dolly the sheep (born 5 July 1996, Roslin Institute, Edinburgh) was the first mammal produced by somatic-cell nuclear transfer (SCNT) from an adult body cell. The table below summarises the published efficiency of that experiment. 9 marks
| Stage of the Dolly experiment | Number | % of previous stage |
|---|---|---|
| Reconstructed embryos created (donor nucleus fused with enucleated egg) | 277 | |
| Embryos that developed enough to be implanted into surrogate ewes | 29 | ≈ 10.5% |
| Pregnancies established | 13 | ≈ 44.8% of implanted |
| Live lambs born | 1 (Dolly) | ≈ 7.7% of pregnancies |
| Live lambs from total reconstructed embryos | 1 | ≈ 0.36% |
Data: Wilmut et al. (1997), Nature 385: 810–813 ("Viable offspring derived from fetal and adult mammalian cell nuclei").
1.1 Express the overall success rate of the Dolly experiment as a ratio (embryos : live lambs) and as a percentage. 2 marks
1.2 Identify two stages of the workflow where the largest drop-offs occurred, and suggest one biological reason for each loss. 4 marks
1.3 A newspaper article in 1997 claimed: "Dolly proves that cloning is now an efficient way to copy mammals." Use the data to evaluate that claim in 2–3 sentences. 3 marks
2. SCNT efficiency across species, bar graph
The figure below shows the published live-birth efficiency of somatic-cell nuclear transfer for several mammalian species, expressed as live births per 100 reconstructed embryos. 6 marks
Indicative figures compiled from published SCNT studies (after Wilmut et al. 1997; Wakayama et al. 1998; Polejaeva et al. 2000; Lee et al. 2005; Liu et al. 2018). Rounded for teaching use.
2.1 Identify the species with the highest and lowest live-birth efficiency, and state both values. 2 marks
2.2 Even the highest bar in the graph is only about 2%. Use this to evaluate the lesson's claim that "whole-organism cloning has low efficiency". 2 marks
2.3 A biotech start-up uses this graph to advertise: "Cloning is more reliable in some species, so our service is safe." Identify one weakness in that advertisement. 2 marks
3. The Tasmanian thylacine, should we try to clone an extinct animal?
The thylacine (Thylacinus cynocephalus) was a marsupial carnivore endemic to Tasmania. The last known animal died at Hobart Zoo in 1936. Read the timeline, then answer the questions. 9 marks
1936 Last confirmed thylacine ("Benjamin") dies in Hobart Zoo. The species is later declared extinct.
1999 Australian Museum (Sydney) launches a project to clone the thylacine using DNA recovered from a preserved 1866 pup specimen.
2002 Project reports successful PCR amplification of fragments of thylacine DNA, but the DNA is heavily degraded.
2005 Australian Museum officially abandons the cloning project, citing degraded DNA and the absence of a viable cell.
2008 Pask et al. (PLOS ONE) insert a thylacine gene (Col2a1 enhancer) into mouse embryos using gene-cloning techniques; the gene is shown to be functional in mouse cartilage.
2022 University of Melbourne + Colossal Biosciences announce a new project aiming to use gene-editing of a related living marsupial (fat-tailed dunnart) plus an artificial surrogate, NOT classical SCNT, to attempt "de-extinction".
3.1 Explain why the 1999–2005 Australian Museum project was unable to use somatic-cell nuclear transfer to clone the thylacine, even with thylacine DNA available. 3 marks
3.2 The 2008 Pask et al. study used gene cloning, not whole-organism cloning. Explain why gene cloning was a more achievable goal for that team, with reference to the lesson's distinction between the two technologies. 3 marks
3.3 Even if the 2022 project produces a live animal, why would the lesson's "phenotype is not guaranteed" warning still apply? Give one specific biological reason. 3 marks
4. Apply gene cloning to a real scenario, human insulin
Before the 1980s, insulin used to treat diabetes was extracted from the pancreases of pigs and cattle, with about 1 kg of insulin requiring around 8,000 kg of pancreas tissue. In 1982 Eli Lilly's "Humulin" became the first marketed therapeutic produced by gene cloning: the human insulin gene was inserted into a bacterial plasmid, transferred into E. coli host cells, and the cells cultured to produce human insulin in industrial fermenters. 6 marks
4.1 Identify the vector and the host cell used in the Humulin process, and state the role of each. 2 marks
4.2 Explain two reasons why gene cloning is more effective than the pre-1980s animal-extraction method for producing insulin. 2 marks
4.3 Using the Humulin example, justify the lesson's claim that "gene cloning's effectiveness is often easier to justify than whole-organism cloning's effectiveness" in 1–2 sentences. 2 marks
Q1.1, Overall success rate of Dolly (2 marks)
Ratio is approximately 277 : 1 reconstructed embryos to live lambs [1]. As a percentage that is 1/277 ≈ 0.36% (accept 0.3–0.4%) [1].
Q1.2, Largest drop-offs (4 marks)
Largest losses occurred (i) between reconstructed embryos (277) and embryos suitable for implantation (29), a ~89.5% loss [1]; and (ii) between pregnancies established (13) and live lambs (1), a ~92% loss [1]. Biological reasons (any one each, 1+1 marks): embryos fail to reprogram the donor nucleus and so stall in early cleavage; many implanted embryos fail to develop normally because of epigenetic abnormalities in the donor DNA; many pregnancies miscarry due to placental defects ("large offspring syndrome"); developmental failures during gestation due to inappropriate gene expression from the somatic donor nucleus.
Q1.3, Evaluate the 1997 newspaper claim (3 marks)
The claim is not supported by the data [1]. Producing one live lamb from 277 reconstructed embryos (≈0.36% success) is the opposite of an efficient process, most attempts failed at one of several stages, including over 90% loss between pregnancy and live birth [1]. The data shows that whole-organism cloning is biologically possible, but its effectiveness as a routine "way to copy mammals" was extremely low, exactly the limitation the lesson highlights in Card 2 [1].
Q2.1, Highest and lowest SCNT efficiency (2 marks)
Highest: cow ≈ 2.0% [1]. Lowest: sheep ≈ 0.4% (Dolly's species) [1]. (Macaque at 0.6% is also accepted as second-lowest reasoning.)
Q2.2, "Whole-organism cloning has low efficiency" (2 marks)
The claim is strongly supported [1]. Even the best-performing species (cow, dog) require ≈50 reconstructed embryos to produce one live birth, and most species lose 98–99% of attempts somewhere in the workflow, well outside what is typically called an "efficient" biological process [1].
Q2.3, Weakness in the start-up's advertisement (2 marks)
The advertisement conflates "more reliable than other species" with "reliable" [1]. Even cow SCNT at ~2% still means ~98% of attempts fail; the graph does not say anything about the health, longevity or phenotype of the surviving clones, so "safe" cannot be justified from these data [1].
Q3.1, Why SCNT failed for the thylacine project (3 marks)
SCNT requires a viable donor cell with an intact nucleus that can be reprogrammed inside the enucleated egg [1]. The thylacine DNA available in 1999 came from a 19th-century pup preserved in ethanol, the DNA was fragmented and degraded, with no living cell or intact nucleus to transfer [1]. PCR could amplify short fragments, but you cannot reconstruct a whole, functioning, ordered genome to drive embryonic development from those fragments, so SCNT was not a feasible technique with the available material [1].
Q3.2, Why gene cloning was achievable (3 marks)
Gene cloning targets a single selected DNA sequence and inserts it into a vector for replication in a host cell, the goal is narrow and the technique tolerates short DNA fragments [1]. Pask et al. only needed one functional region of one thylacine gene (the Col2a1 enhancer), which could be amplified from degraded DNA and ligated into a vector [1]. Whole-organism cloning would have required reconstructing the entire genome inside a viable egg, a much harder organism-level goal that was impossible with the available degraded material, which is why the team chose the gene-level approach [1].
Q3.3, "Phenotype not guaranteed" applied to the 2022 project (3 marks)
Even if gene-edited dunnart cells produce a live "thylacine-like" animal, its phenotype is not guaranteed to match a real thylacine [1]. The marsupial surrogate provides a different uterine environment from a real thylacine mother; epigenetic reprogramming of the edited nucleus may be incomplete; and many phenotypic traits (size, coat pattern, behaviour, gut microbiome) depend on developmental and environmental conditions, not just nuclear DNA [1]. So we may produce an organism with thylacine-like DNA at some loci, but not a true ecological or behavioural restoration of the species, exactly the lesson's warning that genotype copying does not guarantee identical phenotype [1].
Q4.1, Vector and host in Humulin (2 marks)
Vector: bacterial plasmid, carries the inserted human insulin gene into the host and replicates inside it [1]. Host cell: E. coli receives the recombinant plasmid, divides repeatedly to produce many copies of the gene, and expresses the gene to produce human insulin protein, which is harvested from culture [1].
Q4.2, Two reasons gene cloning beats animal extraction (2 marks)
Any two of: (a) the insulin produced is identical to the human protein rather than slightly different pig/cow insulin, reducing immune reactions in patients; (b) production scales by simply growing more bacteria in fermenters rather than slaughtering thousands of animals; (c) it is far cheaper per kilogram and not limited by livestock supply; (d) the source is safer, no risk of pathogen contamination from animal pancreas tissue [1 each, max 2].
Q4.3, Gene cloning's effectiveness vs whole-organism cloning's (2 marks)
The Humulin process has a narrow, measurable goal (produce many copies of the human insulin gene and its protein) and bacterial fermentation routinely achieves this at industrial scale [1]. Whole-organism cloning has a much broader goal, produce a developmentally viable organism, and the Dolly data (≈0.36% success) shows that achieving that goal at acceptable rates is far harder. The narrower, well-defined goal is exactly why gene cloning's effectiveness is easier to justify [1].