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Molecular Biology Final Exam (150 Points – Q1 and Q2 are worth 30pts and Q3 and 4 is at 40 points) Techniques/Controls/Discussion/Context all critical

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Molecular Biology Final Exam (150 Points – Q1 and Q2 are worth 30pts and Q3 and 4 is at 40 points)

Techniques/Controls/Discussion/Context all critical. Expecting 20 plus pages with flow diagrams!

1) Your lab has been studying a new oncogene ZOM that has been found to bind another protein BIE in an initial 2-hybrid screening. Techniques/Controls/Discussion/Context

A) Discuss the 2-Hybrid methods with the controls needed to confirm the ZOM-BIE protein complex.

B) Discuss two other protein-protein technologies with controls that should be used to verify this protein interaction.

C) Design a series of site-directed mutagenesis experiments to test the protein interaction domains in the two proteins with controls.

2) Another Lab has found that the BIE protein is also an oncogene as well. You then propose in your lab to test the expression levels of both ZOM and BIE in a normal series of lung, brain, liver, and kidney cell lines and cancer cell lines derived from the same tissues. Techniques/Controls/Discussion/Context

A) You first develop a microarray to test for the genes encoding the ZOM and BIE proteins in all eight cell lines. Discuss the practical use of the microarrays as well controls to establish the expression levels on these genes.

B) You also have antibodies to the ZOM and BIE proteins and proceed to develop a protein array analysis as well. Discuss experiments and controls.

C) Finally, discuss RNA SEQ as a method to determine the level of mRNA, alternative splicing, and cell distribution with controls.

3) As you have studied the ZOM and BIE, it turns out they are Signaling Kinases connected to the MYC oncogene pathway. Techniques/Controls/Discussion/Context

A) Discuss the Myc/Max/Mad etc. Pathway in general and how would you determine where the ZOM and BIE protein kinases are in that cascade, and how would you test that hypothesis.

B) In your experiments with the Myc/Max/Mad etc. The pathway you find that ZOM and BIE inter in RAS Transcription Factor network as well. The discussion that transcriptional network’s structure and function. How would you determine the ZOM and BIE interactions with RAS/RAF etc.?

4) Finally, in the cancer cell lines ZOM and BIE function in regulating the CDC 25 Phosphatase and Influence the RB protein phosphorylation state and its metabolism within the Cell Cycle. Techniques/Controls/Discussion/Context

A) Discuss the cell cycle checkpoints, CDC 25 and its structure and function, MPF, and RB.

B) Outline and discuss experiments with controls to confirm ZOM and BIE interaction in the Cell Cycle