LABORATORY SCHEDULE - Fall 2007

8:30

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8:45

Laboratory Course Objectives (Sharon Norton)

8:45

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9:30

Lecture: Carbonic Anhydrase Model (Dr. David Silverman) 

9:30

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10:30

Lecture: Introduction to Protein Structure (Dr. Sophie Alverez)

10:45

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12:00

Exp. 2: Lysis of E. coli

Exp. 4: Affinity purification up to sample incubation.

12:00

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12:45

Lunch (on your own)

12:45

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1:45

Lab:  Bioinformatics I: I ntroduction to Web-based tools (Dr. Fahong Yu)

1:45

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2:45

Exp 3A,B,C: SDS-PAGE Analysis of bacterial lysate fractions.

2:45

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4:15

Exp. 4 cont. and Exp. 5: Affinity purification, de-salting and concentration by ultrafiltration

4:15

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5:00

Exp. 3D: gel staining and destaining overnight.

8:30

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8:40

Lab review: What’s next?

8:40

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9:30

Exp. 6: Protein Concentration Assay.

9:30

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10:15

Exp. 8:  Minigel apparatus. Pour separating gel. Tris-glycine system.

10:15

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11:15

Lecture: Theory of Electrophoresis (Dr. Bill Farmerie)

11:15

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11:45

Exp. 8: Pour stacking gel

11:45

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12:45

Lunch

12:45

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1:45

Exp. 6:  Protein concentration determination (Excel help, Dr. Fahong Yu)

1:45

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2:45

Exp. 8: Prepare, load samples, electrophorese.

2:45

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3:45

Exp. 9A:  Set-up protein transfer.

3:45

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4:15

Exp. 3E: Gel Drying

4:15

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5:00

Exp 9B,C: Stop Western Transfer /Block O/N.

8:30

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9:00

Exp. 9C: Add 1^o Ab to immunoblots.

9:00

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10:00

Lecture:  Winning Westerns (Linda Green).

10:00

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10:30

Exp. 9C: Wash Nitrocellulose membrane/Add 2^o Ab.

10:30

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12:00

Lab lecture: Bioinformatics II: Introduction to Bioinformatics Tools

                     Bioinformatics III: Assignment of projects  (Dr. Fahong Yu).

12:00

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1:00

Lunch

1:00

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2:00

Exp 9C: Finish Western Blots (wash, color substrate addition).

2:00

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3:00

Lecture: Computer Modeling of Proteins (Dr. Ben Dunn)

300

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3:30

Exp. 11B:  Start first dimension of Two-D electrophoresis.

4:00

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5:00

Graduate Students Reception: ARB Founders Gallery

1:00

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2:00

Lecture: Biological Mass Spec Overview and Sample Preparation (Dr. Sophie Alverez)

2:00

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5:30

Exp 12A: Reduction/ Alkylation and trypsin digestion of glycoprotein.

8:30

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9:30

Seminar: Animal Awareness (Charles Mason)

9:30

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10:15

Exp 12C: Preparation of Zip tips and spotting the plate.

10:15

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11:00

Exp. 11C,D: Equilibration of IEF strips and begin 2^o dimension

11:00

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12:00

Lecture: 2D Gel Electrophoresis (Dr. Sophie Alvarez).

12:00

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12:30

Exp. 11D, E:  Disassemble two-dimensional gel (2^o dimension), stain.

12:30

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1:30

Lunch

1:30

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2:00

Exp 11E: Destaining of 2D gel.

2:00

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2:45

Lecture:  Immunocytochemical detection of antigens. (Steve McClellen)

2:45

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3:45

Demo: Viewing of fluorescently and colormetrically immunostained slides.

3:45

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4:30

Lecture: Flow Cytometry (Neal Benson).

8:30

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8:40

Lab Review: What’s next?

8:40

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9:15

Exp. 14A: RNA Isolation and spectrophotometric analysis.

9:15

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10:15

Exp. 14B: Cleaning of apparatus and pouring of gel.

                 Preparing the samples and loading the gel.

10:15

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11:45

Lecture and Demo: RNA Isolation and QC: Bioanalyzer (Dr. Gigi Ostrow)

11:45

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12:00

Exp. 14: Photograph RNA gel

12:00

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1:00

Lunch

1:00

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2:00

Lecture: Bioinformatics IV-Designing your own primers for the RT-PCR of mouse CAII (Dr. Fahong Yu).

2:00

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3:00

Lab: Designing your own primers and optimization experiment for the RT-PCR of mouse CAII

3:00

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4:00

Exp. 15: Set up reverse transcription and PCR reaction.

4:00

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5:00

Lecture: PCR Theory (Dr. Bill Farmerie )

8:30

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8:40

Lab review: What’s next?

8:40

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9:15

Exp.16: Gel electrophoresis of RT-PCR samples.

9:15

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10:15

Lecture: Gene Cloning (Dr. David Moraga)

10:15

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12:15

Exp. 16A,B,C: Excise bands. Purify DNA from gel slice. Pour gel.

12:15

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1:00

Lunch

1:00

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1:30

Exp. 16D: Start TOPO ligation system (bacterial transformation).

1:30

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2:30

Exp. 12: Mass Spectrometry analysis: Database searching demo and computer time (guided by Scott McClung).

2:30

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3:15

Exp 16D: Spread X-gal on agar plates. Plate cells.

3:15

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4:15

Lecture: Practical Aspects of DNA Sequencing (Dr. Ernie Almira).

11:00

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12:00

Exp. 17A: Begin gDNA isolation (up to 100% ethanol precipitation).

12:00

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1:00

Lunch

1:00

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2:00

Exp. 17A: Finish DNA isolation, pour gel.

2:00

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2:45

Exp 17B: Agarose gel analysis of genomic DNA

2:45

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3:45

Lecture: Bioinformatics V - In Silico cloning and screening (Dr. Fahong Yu).

3:45

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4:15

ExpB,C: Stain, take picture, quantitate genomic DNA.

4:15

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4:45

Exp 17C: Set-up incremental digestions

4:45

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5:00

Exp. 16E: Begin overnight cultures from individual bacterial colonies.

11:00

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11:30

Exp. 17D: Agarose electrophoresis of gDNA for Southerns.

11:00

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12:00

Exp. 16E2-E3, F: Plasmid miniprep, set up restriction enzyme digestions.

12:00

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1:00

Lunch

1:00

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1:30

Exp. 17D: Lab assistant will take photograph of digested DNA gels.

1:30

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2:30

Exp. 17E: Set up Southern Transfer

2:30

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3:00

Exp. 16G: Gel electrophoresis.

3:00

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4:00

Lecture: Real Time PCR (TBA).

4:00

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4:30

Exp 16G: Photograph gel.

11:00

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11:30

Exp. 17E-17F1: Dismantle Southern, UV-crosslink, begin prehyb.

11:30

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12:30