Table of contents

1. Introduction to Biology2h 42m
2. Chemistry3h 37m
3. Water1h 26m
4. Biomolecules2h 23m
5. Cell Components2h 26m
6. The Membrane2h 31m
7. Energy and Metabolism2h 0m
8. Respiration2h 40m
9. Photosynthesis2h 49m
10. Cell Signaling59m
11. Cell Division2h 47m
12. Meiosis2h 0m
13. Mendelian Genetics4h 44m
14. DNA Synthesis2h 27m
15. Gene Expression3h 6m
16. Regulation of Expression3h 31m
17. Viruses37m
- Viruses
  37m
18. Biotechnology2h 58m
19. Genomics17m
- Genomes
  17m
20. Development1h 5m
21. Evolution3h 1m
22. Evolution of Populations3h 53m
23. Speciation1h 37m
24. History of Life on Earth2h 6m
25. Phylogeny2h 31m
26. Prokaryotes4h 59m
27. Protists1h 12m
28. Plants1h 22m
29. Fungi36m
- Fungi
  19m
- Fungi Reproduction
  17m
30. Overview of Animals34m
- Overview of Animals
  34m
31. Invertebrates1h 2m
32. Vertebrates50m
33. Plant Anatomy1h 3m
34. Vascular Plant Transport1h 2m
- Water Potential
  1h 2m
35. Soil37m
- Soil and Nutrients
  20m
- Nitrogen Fixation
  16m
36. Plant Reproduction47m
- Flowers
  33m
- Seeds
  14m
37. Plant Sensation and Response1h 9m
38. Animal Form and Function1h 19m
39. Digestive System1h 10m
- Digestion
  1h 3m
- Blood Sugar Homeostasis
  7m
40. Circulatory System1h 49m
41. Immune System1h 12m
42. Osmoregulation and Excretion50m
- Osmoregulation and Excretion
  50m
43. Endocrine System1h 4m
- Endocrine System
  1h 4m
44. Animal Reproduction1h 2m
- Animal Reproduction
  1h 2m
45. Nervous System1h 55m
- Neurons and Action Potentials
  1h 8m
- Central and Peripheral Nervous System
  46m
46. Sensory Systems46m
- Sensory System
  46m
47. Muscle Systems23m
- Musculoskeletal System
  23m
48. Ecology3h 11m
49. Animal Behavior28m
- Animal Behavior
  28m
50. Population Ecology3h 41m
51. Community Ecology2h 46m
52. Ecosystems2h 36m
53. Conservation Biology24m
- Conservation Biology
  24m

18. Biotechnology

Steps to DNA Cloning

General Biology

18. Biotechnology

Steps to DNA Cloning

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2:18 minutes

Problem 11

Textbook Question

Explain how you might engineer E. coli to produce human growth hormone (HGH) using the following: E. coli containing a plasmid, DNA carrying the gene for HGH, DNA ligase, a restriction enzyme, equipment for manipulating and growing bacteria, a method for extracting and purifying the hormone, and an appropriate DNA probe. (Assume that the human HGH gene lacks introns.)

Verified step by step guidance

Step 1: Use a restriction enzyme to cut the plasmid DNA from E. coli and the DNA carrying the human growth hormone (HGH) gene. Restriction enzymes recognize specific sequences in DNA and create cuts, often leaving 'sticky ends' that can facilitate the joining of DNA fragments.

Step 2: Combine the cut plasmid DNA and the HGH gene DNA. The sticky ends created by the restriction enzyme will allow the two DNA fragments to align and bond. Use DNA ligase to seal the sugar-phosphate backbone, creating a recombinant plasmid that contains the HGH gene.

Step 3: Introduce the recombinant plasmid into E. coli cells through a process called transformation. This can be achieved by making the bacterial cells competent (e.g., using heat shock or electroporation) to take up the plasmid DNA.

Step 4: Grow the transformed E. coli cells in a nutrient-rich medium. The bacteria will replicate, and as they do, they will also replicate the recombinant plasmid, leading to the production of human growth hormone. Ensure the plasmid includes a promoter sequence that allows the HGH gene to be expressed in the bacterial cells.

Step 5: Extract and purify the human growth hormone from the bacterial culture. Use a method such as affinity chromatography to isolate the hormone. To confirm the presence of the HGH gene in the plasmid, use an appropriate DNA probe that hybridizes specifically to the HGH gene sequence.

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Key Concepts

Here are the essential concepts you must grasp in order to answer the question correctly.

Recombinant DNA Technology

Recombinant DNA technology involves combining DNA from different sources to create new genetic combinations. In this context, it allows for the insertion of the human growth hormone (HGH) gene into a plasmid, which can then be introduced into E. coli. This process enables the bacteria to express the HGH protein, which can be harvested for therapeutic use.

Plasmids and Cloning Vectors

Plasmids are small, circular DNA molecules that can replicate independently within a bacterial cell. They are commonly used as cloning vectors to carry foreign genes, such as the HGH gene, into host organisms like E. coli. The plasmid must contain a promoter for gene expression and a selectable marker to identify successfully transformed cells.

Gene Expression and Protein Purification

Gene expression refers to the process by which the information in a gene is used to synthesize a functional protein. After E. coli has been transformed with the plasmid containing the HGH gene, the bacteria will produce HGH. The final step involves extracting and purifying the hormone from the bacterial culture, which may include techniques like affinity chromatography to isolate the HGH protein from other cellular components.

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Related Practice

Textbook Question

Which of the following sequences in double-stranded DNA is most likely to be recognized as a cutting site for a restriction enzyme?

a. AAGG TTCC

b. GGCC CCGG

c. ACCA TGGT

d. AAAA TTTT

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Textbook Question

A biochemist hopes to find a gene in human cells that codes for an important blood-clotting protein. She knows that the nucleotide sequence of a small part of the blood-clotting gene is CTGGACTGACA. Briefly outline a possible method she might use to isolate the desired gene.

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Textbook Question

A biologist isolated a gene from a human cell, inserted it into a plasmid, and inserted the plasmid into a bacterium. The bacterium made a new protein, but it was nothing like the protein normally produced in a human cell. Why? (Explain your answer.)

a. The bacterium had undergone transformation.

b. The gene did not have sticky ends.

c. The human gene contained introns.

d. The gene was not synthesized from scratch.

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Textbook Question

Potato blight causes potato plants to shrivel and rot. The disease is caused by the pathogen Phytophthora infestans, infamous for its role in Ireland's Great Potato Famine in the mid-1840s. The disease can devastate crops during wet weather, sometimes leading to total crop loss. Researchers aim to use recombinant DNA methods to transfer blight resistance genes from resistant varieties into susceptible varieties of potato.Explain how restriction endonucleases and DNA ligase could be used to insert a potato blight resistance gene into a plasmid.

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Textbook Question

Potato blight causes potato plants to shrivel and rot. The disease is caused by the pathogen Phytophthora infestans, infamous for its role in Ireland's Great Potato Famine in the mid-1840s. The disease can devastate crops during wet weather, sometimes leading to total crop loss. Researchers aim to use recombinant DNA methods to transfer blight resistance genes from resistant varieties into susceptible varieties of potato. Explain how restriction endonucleases and DNA ligase could be used to insert a potato blight resistance gene into a plasmid.

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Textbook Question

There are difficulties inherent in testing hypotheses involving human health. Imagine that you have developed a potential vaccine against AIDS. Given the nature of the disease, all testing must be conducted in humans. Can you design an ethical but effective experiment?

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Multiple Choice

Human DNA cut with restriction enzyme A can be joined to:

Each restriction enzyme has a specific sequence of nucleotides where it cuts the DNA. These sequences of DNA are unique to each restriction enzyme and are known as:

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