Which one of the following best describes what can be done by variations on the CRISPR/Cas9 DNA editing system?

The lac

operon is used to regulate genes in prokaryotic cells. Which one of the following

statements best describes the mechanism of the lac operon in

What

regulatory hurdle must biosimilar manufacturers overcome that is not required

for generic drugs?

(i)   Name the overall process represented by the diagram below. (1 mark) 

(ii)  With reference to the diagram, explain your rationale for your answer in part (i) (3 marks) 

(iii) In an experiment, you added an inhibitor to the component labelled ‘B’.  What will the effect of this inhibitor have on the process shown below? (1 mark) 

[1 + 3 + 1 = 5 marks]

Look at the series of images below depicting an essential cellular process:

In your own words, describe what is being represented by the images, including the name of the process represented and the key components involved.

In Question 1, part C, you chose a specific protein tag or fusion protein. 

(i)   Explain how you would purify the protein expressed from this plasmid.

(ii)  How would you verify the purity of the product that you produced?

Once inserted into the plasmid shown in Question 1, the gene 3021 is controlled by the lac operon. 

(i)   How does the lac operon exert transcriptional control?

(ii)  You plan to grow the bacteria transformed with pBPS-3021 in the presence of glucose initially and then, once all of the glucose is exhausted, add lactose to the medium.  Is this necessary?  Give a brief rationale for your response.

(10 marks)

The plasmid shown in Q1 is a piece of DNA.  Explain how this can be used to produce a protein?

You wish to use traditional (PCR and restriction enzyme-based) cloning techniques to clone your gene of interest (gene 3021) into the plasmid vector pBPS (to make the recombinant plasmid pBPS-3021).  You wish to express the gene product as a fusion protein in E. coli. 

For each of the plasmid features (A-E), as shown on the diagram above, answer the following questions:

A.  What is the purpose of the promoter and does it need to be before (upstream of) the MCS as shown in the diagram?

B.  The gene of interest will be inserted into the MCS, briefly explain how this will be achieved.

C.  Name one "tag" or fusion protein commonly used in plasmid expression vectors, and, based on the figure provided, briefly explain whether or not you will need to insert the 3021 gene in-frame with this tag.

D.  What is the purpose of the origin of replication?  Do high-copy plasmids have multiple ori's?

E.  Name one selectable marker commonly used in plasmid expression vectors and briefly explain its purpose.

(2 marks each)