Molecular Cloning: Cell Bio Lab.

Molecular Cloning: Cell Bio Lab.

Form #: 23

Cell Bio Lab Instructions Sheet

Please use the information in this Instructions Sheet, that you have been uniquely assigned, to complete your Answer Sheet. Be sure to record your Name, Lab section, and Form # on your answer sheet word doc (available for download in the assignment instructions in Canvas). Work on your own and be sure short answer written responses are all typed and paraphrased in your own words to receive credit.

1. How do you anticipate production of recombinant DHFR may help identify new anti-cancer therapeutics? In your answer, include the relevance of DHFR function to cancer and how the recombinant protein may be useful for future therapeutic research. (3pt)

2. Identify one protein with a normal, endogenous biological function in nature that we have used in the lab to produce recombinant DHFR. Where does this protein come from (biological source/organism where it is naturally produced) and what is the protein’s normal, endogenous function? How did we specifically make use of this protein in the lab for our project, to produce recombinant DHFR? (6pt)

3. Which protein tag did we use to increase recombinant DHFR solubility? (1pt)

4. What would happen if you accidentally used the bacteria that we use for molecular cloning steps instead of the genetically engineered BL21 (DE3) cells for recombinant DHFR protein expression? Do you anticipate the amount of recombinant protein produced would be impacted and if so, why? Please be specific and include molecular level detail in your answer. (3pt)

5. Imagine you followed all steps correctly and found that the amount of recombinant DHFR produced through our method was low. What is one thing you could do to try to increase the protein yield (amount produced) and why do you anticipate it to increase protein production? Please only include and discuss one answer here. (4pt)

6. How would you make 50uL 1:10, 1:100, and 1:1000 serial fold dilutions from a starting 1:1 solution? Please complete the following table. (3pt)

*Notation here denotes part:whole

7. Briefly explain how you could use the color of the Coomassie assay sample solution to determine if an unknown sample needs to be diluted in order to measure the sample’s concentration. (2pt)

8. List and describe one source of variation in the hands-on steps for Coomassie assay standard curve generation. How will you know if your standard curve is significantly impacted by experimental error due to this potential source of variation? (3pt)

Name:_________________________             Lab Section:______                                                         Form #:____

Cell Bio Lab Answer Sheet

. Please use the resources/tools from lab activities covered so far this semester. Be sure that all short answer writing is typed and paraphrased in your own words, do not copy text directly from another source. Please complete, save, and upload this word doc answer sheet to Canvas, Turnitin enabled. (25pt)

1. (3pt):

2. Protein (1pt): ____________

Biological source/organism (1pt):_________________

Normal (endogenous) function (2pt):

Specific role in our lab project (2pt):

3. Tag (1pt): _________

4. (3pt):

5. Include and discuss one answer (4pt):

6. (3pt):

7. Briefly explain (2pt):

8. Source of variation (1pt):_________

Describe (1pt):

How to know if curve is impacted (1pt):

Molecular Cloning: Cell Bio Lab.