1. Biology 22 Molecular Laboratory Report 1. Bacterial Transformation 2. Plasmid Isolation 3. RFLP Analysis

2. Molecular laboratory report Results are prepared outside of class and handed in on November 3 rd. The Discussion is written in class on November 5 th ; your results will be returned to you to assist you on Nov. 5 th in the writing of your Discussion. No other outside material or electronic devices can be used for writing the Discussion. You have the full class period to write your Discussion. The length should not me more than 6 single-sided sheets and no less than 4 single sided sheets. Paper will be provided.

3. Guidelines for the Results  Different science courses and scientific journals use varied approaches for the presentation of experimental Results but this molecular laboratory report will have only the presentation of the data but not a discussion of the data within the Results section.  The next few slides provide guidelines for preparing and submitting your Results.  The legends are not where data are discussed; simply present what the table or figure presents.  Again, the Result section is not the Discussion and must not include a discussion of the results.  Onto the specifics of the Results…

4. Guidelines for the Results  Include a Title Page  Provide a descriptive title  Your name  Must include group numbers (not individual lab partners) for each experiment  Results  All tables have titles and numbers above table  All figures have legends and numbers below figure  (Discussion: will be written in class on Nov. 5 th)

5. What your Result Section must include: A Table that includes the Transformation results. The Table should be numbered and have a Title. The Title goes above the Table. The Calculation of your Transformation efficiency –If you did not obtain any transformation; this should be mentioned in your discussion; however since everyone observed results from a group who did have results from this experiment it is acceptable to use this for your transformation efficiency. A Photograph with a legend or caption of the isolated plasmid. The caption and figure number both are placed below the figure. A Photograph with a legend or caption of the RFLP. The legend or caption should be numbered and go beneath the figure. A graph of the Standard Curve with a legend or caption. A Table (with title) that summarizes the sizes and distances migrated for the ladder A Table (with title) that summarizes the sizes and distances migrated of the plasmid fragments from each appropriate lane Individual Maps for each restriction digest. Consider these maps collectively as a single figure so include a caption with this figure. Maps are not made for the ladder or for the uncut plasmid

6. Results: Transformation Table: results of 3 agar plates: number and color of colonies Calculation: of the Transformation efficiency

7. Results: plasmid isolation  The photograph of the plasmid isolation.  You should include a numbered legend or caption to this photograph. The legend includes: what samples are present on the gel, their “lane” number, and the type and percent gel.  Be sure and number the lanes of the gel.  Note: The gels were a 1.5% agarose

8. Results  A photograph of the RFLP analysis with figure legend or caption and number; the legend should be placed beneath the photograph.  Be sure and number the lanes of the gel.

9. Results  Standard curve Distance migrated (cm) DNA bp

10. Results  A separate table that shows the size (base pairs) of fragments versus distance migrated of your ladder (known) fragments. You may not be able to identify all 9 fragments of your ladder but you should be able to identify approximately 5 to 6 (or more) of these fragments.  A table that shows the size (base pairs) versus distance migrated of your unknown fragments

11. Provide MAP Results maps for each of the restriction digests.

12. Examples of MAP Results  These are just examples!  You will have third map with both enzymes and various sizes shown. Total Size: 7000 bp Lane 3 Alu1 Lane 4 Total size 7050 bp Hae 4500 2550

13. Number of Maps Lane 3 Lane 4 Lane 5 Map 1 Map 2 Map 3

14. A Discussion is a logical & creative story based on your data. Lastly, try to enjoy yourself as you put the pieces together

15. Discussion written in class on Nov. 5 th On the next couple of slides there are some questions to help guide you think about the Discussion section of your molecular lab report; however you cannot bring in a pre-written Discussion November 5 th. When you do write your Discussion in class it should not be presented in a “question and answer” format rather you should present an organized essay for your Discussion. I also encourage you to think of other concepts that you might want to address regarding a discussion of your results.

16. Concepts/question to address in your discussion: Transformation 1. What did the results from the transformation show about how the plasmid altered phenotype? 2. What reason(s) would you expect for the fact that transformation efficiency is often times quite low? 3. What is the mechanism for the transformation? 4. Why are ampicillin and X-gal included on the agar plates? 5. What is the purpose for including control plates.

17. Discussion Plasmid Isolation 6. How was it possible to separate cellular proteins from nucleic acids? 7. Why did the photograph show more than one band for the plasmid isolates? 8. How did you know if RNA was present and how could it be removed?

18. Discussion RFLP 9. How do restriction enzymes create unique patterns? 10. Which restriction enzymes are unique enzymes? 11. Which enzyme might be used for a cloning experiment to “insert” a piece of DNA into the plasmid? 12. What are some other possible applications for the RFLP technology other than for simply mapping a plasmid?

19. Discussion  Do NOT think of your discussion as a “Q and A” section.  The questions are only meant to help you address and think about important concepts of each experiment.  I will not be answering these questions since they were all covered in lecture and/or lab. It is really a way to learn and review the material as you think about the data.

20. What you should avoid:  The discussion is not a description of the “materials and methods” for example the discussion does not state such details:  As 200 ul of cells were added to agar plates  As a gel box was filled with buffer  As a camera was used to take a picture  Or any other details of how an experiment was performed are not included in the discussion.

21. The Do Not’s Do NOT make the focus of the discussion a list of errors. Do NOT simply state the data cannot be interpreted because of errors—this is just not true! An effort has been made to see that you have access to photos and data that allow you to meaningfully interpret your experiments. If you had a specific problem with an experiment this should be mentioned in your report but then be sure to discuss the data made available to you. Do NOT copy out of the lab manual (or any other source). Do Not

22. The End