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review questions
1. What DNA sequence elements are required for expression of a cloned gene in a prokaryotic host?
2. What is a strong promoter? Why is a strong promoter not always desirable for expression of a cloned gene?
3. What is a regulatable promoter? How is the E. coli lac promoter used to regulate the expression of a clone gene?
4. The promoter for gene 10 of the E. coli bacteriophage T7 is an example of a strong promoter. How is it used to express a cloned gene?
5. Why is codon optimization often required for production of high levels of a recombinant protein?
6. What are inclusion bodies, and how can their formation be avoided?
7. How can a protein of interest be engineered to be secreted to the medium by E. coli?
8. Discuss some strategies to purify a recombinant protein produced in a prokaryotic host. Consider that a protein may be used as a human therapeutic agent.
9. Why is it sometimes advantageous to integrate a target gene into the chromosomal DNA of a prokaryotic host? How might this be achieved?
10. During the course of integrating a target gene into the chromosomal DNA of the host bacterium, a marker gene may also be inserted into the chromosomal DNA. What strategy could be used to excise only the marker gene?
11. What are the major posttranslational modifications of eukaryotic proteins in the endoplasmic reticulum and Golgi apparatus?
12. Describe the features of a eukaryotic expression vector.
13. What criteria are used to decide if a particular recombinant protein should be produced in