Chem 110L
Introductory Biochemistry Laboratory
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Chem 110L |
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Dr. Kalju Kahn
Office: 2623 PSB-N
Office hours: Tuesday 12:00-1:00 PM and by appointment
Phone: 893-6157
E-mail: kalju@chem.ucsb.edu
Web site: http://www.chem.ucsb.edu/~kalju
Section 1 (MW 6-9:50): Clayton Woodcock cwoodcock@chem.ucsb.edu
Office hours: Chem 1148
Section 2 (TR 2-5:50): Kiel Nikolakakis knikolakakis@chem.ucsb.edu
Office hours: LSB 3209
Section 3 (TR 6-9:50): Kota Kaneshige kkaneshige@chem.ucsb.edu
Office hours: Chem 1132
The purpose of Chem 110L is to offer hands-on experience with modern methods of separation, identification, and characterization of biomolecules. The course will strengthen your understanding of material taught in Chem 142A (Biochemistry Lecture). In Chem 110L, you will do experiments with biomolecules such as nucleic acids, proteins, sugars, and lipids. The 1 hour lecture series focuses on principles behind each experiment, and explains instrumental techniques and methods that you will use to accomplish your goals.
Lecture: Mon 3:00-3:50 Place: Phelps Hall 3505 Lab section 1: Mon 6:00-9:50; Wed 6:00-9:50; Lab section 2: Tue 2:00-5:50; Thu 2:00-5:50; Lab section 3: Tue 6:00-9:50; Thu 6:00-9:50; Lab sections are in PSB-N 2619 unless otherwise noted
| Syllabus | General information about the course. | |
| Schedule | Schedule MW6 | |
| Schedule | Schedule for TR2 | |
| Schedule | Schedule for TR6 | |
| Lab Text | Required Theory Manual: Chapter 1 | |
| Lab Text | Required Theory Manual: Chapter 2 | |
| Textbook | Recommended text 1 | Link |
| Textbook | Recommended text 2 | Link |
| Exam | Preparation for the exam: topic list |
Students in the class do not have to purchase the laboratory manual. Each chapter of the lab manual can be downloaded here in the PDF format. Please note that you can follow hyperlinks that are in the PDF files by clicking on the link. Links to external literature sources are given later below.
| Experiments | Download Adobe Acrobat Here | Acrobat |
| Lab 1: | Macromolecular visualization | Tutorial |
| Lab 2: | Operations Manual: molar absorbtivity of urate | |
| Lab 2: | Operations Manual: Mathematica Tutorial | |
| Lab 3: | Optical microscopy | |
| Lab 4: | Thermal denaturation of double-stranded DNA | |
| Lab 4: | Computational Thermochemistry | Link |
| Lab 5: | Agarose gel electrophoresis of DNA isoforms | |
| Lab 6: | Size Exclusion Chromatography | |
| Lab 6: | Bioinformatics: Protein Identification Tutorial | |
| Lab 6: | Bioinformatics: Protein Identification Assignment | |
| Lab 7: | Quantitative Analysis of Ligand–Macromolecular Interactions | |
| Lab 8: | Quantitative enzymatic determination of glucose | |
| Lab 9: | Determination of the iodine value of a lipid by 13C NMR | |
| Lab 10: | Light-induced proton gradient in chloroplast | |
| Lab X: | Isoelectric Focusing Demonstraton |
| Literature | Required or optional reading in PDF | Acrobat |
| General | Operations Manual: operation of pipettors | |
| General | Fitting Models to Biological Data (advanced) | |
| General | Hands-on Start to Mathematica (Video tutorials) | Link |
| Lab 3: | Olympus BX41 User Guide for Chem110L | |
| Lab 3: | Microscopy: Dr. Matsumoto's Presentation Slides (2005) | |
| Lab 3: | Microscopy: Perspective Story "How to Build a Superlens" by David R. Smith | |
| Lab 3: | Microscopy: Fang et. al.: "Sub-Diffraction-Limited Optical Imaging with a Silver Superlens " | |
| Lab 3: | DNA electrophoresis media: review | |
| Lab 4: | Thermal denaturation of double-stranded DNA (advanced) | |
| Lab 6: | "Gel Filtration: Principles and Methods" | |
| Lab 6: | "Supelco LC Media Selection Guide" | |
| Lab 6: | BioRad columns manual | |
| Lab 6: | BioRad Bio-Gel manual | |
| Lab 6: | ToyoPearl Instruction manual | |
| Lab 6: | BioRad EconoPump Manual | |
| Lab 6: | BioRad Fraction Collector Manual | |
| Lab 6: | Multivariable Spectral Analysis | |
| Lab 6: | Importing and Visualization of Spectra with Mathematica | |
| Lab 7: | New methodologies for measuring protein interactions in vivo and in vitro | |
| Lab X: | Electrophoresis: DryStrip Kit Manual | |
| Lab X: | Electrophoresis: Power Supply Manual | |
| Lab X: | Proteomics: Isoelectric Focussing | |
| Lab X: | Proteomics: Protein Electrophoresis, SDS-PAGE | |
| Lab 8: | Current Problems and Potential Techniques in In Vivo Glucose Monitoring | |
| Lab 9: | 13C NMR (Lipids): Lecture Slides | |
| Lab 9: | 13C NMR (Lipids): Spectra and Peak Lists | Folder |
| Lab 10: | Photosynthesis: Colloquium Paper: Hu et al, PNAS 1998 | |
| Lab 10: | Prokaryotioc Photosynthesis: Bryant & Frigaard, 2006 |
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One of the experiments in the course involved demonstration and use of microscopes. The images below show how fluorescence imaging can be used to visualize specific sub-cellular structures. The first image shows microtubules stained with a fluorescent dye; the image was recorded in black and white. The second image shows nuclei stained with a fluorescent compound DAPI; this image was also recorded in black and white but was obtained using excitation light of different wavelength than the microtubule image. To obtain the last image, separate colors were assigned to the two previous images before combining them into one. |
| Microtubules | Nuclei | Combination |
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One of the instruments used was the Olympus BX41 fluorescence microscope in the laboratory of Prof. Stanley Parsons in the Department of Chemistry and Biochemistry. The images below show three areas of the first commercial slide with bovine pulmonary arterial epithelial cells. Each area was visualized by three fluorescent stains. The first stain, DAPI, binds to DNA and makes nuclei fluoresce in blue. The second stain, AlexaFluor 488, has been coupled to phalloidin, which binds to actin, allowing visualization of actin filaments of the cytoskeleton. The third dye is MitoTracker Red, which binds to mitochondria. The fluorescence from the sample was resolved with 100X oil immersion objective and captured with a CCD camera in gray-scale. You can click on each image to download the high-resolution file. |
| Nuclei | Mitochondria | Actin |
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The next set of images shows two areas of a second commercial slide with bovine pulmonary arterial epithelial cells. Each area was visualized by three fluorescent stains. The first stain, DAPI, binds to DNA and makes nuclei fluoresce in blue. The second stain, BODYPY FL, has been coupled to a tubulin-specific antibody molecule, allowing visualization of microtubules. The third dye is phalloidin-conjugated Texas Red-X, which binds to actin. The first set of images represent cells in the prophase of mitosis. The second set shows another cell during early anaphase. The fluorescence from the sample was resolved with 100X oil immersion objective and captured with a CCD camera in gray-scale. You can click on each image to download the high-resolution file. |
| Nuclei | Microtubules | Actin |
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