Chabot College
Course Outline for Biotechnology 50, page 3
Fall 2010
Chabot College Fall 2010
Course Outline for Biotechnology 50
Biotechnology Laboratory Skills II
Catalog Description:
50 – Biotechnology Laboratory Skills II 2 units
Introduces students who are interested in biotechnology, biological sciences, and current industry workers to the advanced laboratory research methods and concepts in biotechnology. Laboratory skills include mastering the tools used in biotechnology such as isolation and quantification of DNA, amplifications with PCR, media preparation and dilution, aseptic technique, and cell culture. Prerequisite: Biotechnology 40 (completed with a grade of “C” or higher). Strongly recommended: Mathematics 54 (completed with a grade of “C” or higher) or appropriate skill level as demonstrated by the mathematics placement test, and eligibility for English 1A. 1 hour lecture, 3 hours laboratory.
[Typical contact hours: lecture 17.5, laboratory 52.5]
Prerequisite Skills:
1. demonstrate appropriate behaviors, teamwork, and proper safety procedures to work in a laboratory environment, including maintaining a professional quality laboratory notebook;
2. demonstrate an understanding of the scientific method, experimental design, data collection, basic statistics, basic laboratory skills, and procedures including the preparation of reagents and other materials;
3. demonstrate the ability to make solutions, reagents, buffers;
4. demonstrate basic concepts and applications of chemistry and biochemistry appropriate for a biotechnology laboratory, with the goal of preparing students to work with basic and sophisticated instrumentation in a biotechnology laboratory, e.g., spectrophotometers, electrophoresis apparatus, pH meters, and chromatographic systems;
5. demonstrate extraction of DNA from E. coli, its characterization and quantification using agarose gel electrophoresis, and UV spectrophotometry;
6. demonstrate expertise with the use of the classical tool-kit of molecular biology such as restriction enzymes, vectors, plasmids, host cells, and expertise of the polymerase chain reaction (PCR) to clone several loci from the human genome;
7. demonstrate the tools of SDS-PAGE and Western Blotting to characterize purified proteins;
8. demonstrate the proper procedures for the aseptic culturing of microorganisms, their preparation for microscopy (e.g., Gram staining), and their use as vectors in recombinant DNA work.
Expected Outcomes for Students:
Upon completion of this course, the student should be able to:
1. demonstrate understanding of SOPs;
2. demonstrate ability to prepare media, gels, buffer solutions and reagents of specific concentrations by using the appropriate stock solutions;
3. isolate and purify DNA from biological systems and plasmid vectors from bacterial for cloning;
4. demonstrate the proper use of spectrophotometry to assess the purity and quantity of DNA that have been isolated, and to determine population density of bacterial cultures;
5. prepare competent cells for transformations with recombinant vectors and calculate transformation efficiency;
6. analyze and interpret results from electrophoretic gels;
7. construct standard curve graphs relating known DNA fragment size in the estimation of size of unknown DNA fragments;
8. construct restriction maps from data obtained from electrophoresis of restriction digests;
9. properly prepare and use selective media (with antibiotics as the selective base) to eliminate nontransformed specimens (level one screening);
10. use the products of PCR as a source of DNA for cloning specific genes from other organisms;
11. design PCR primers on the basis of phylogenetic and nucleotide sequence information gathered from electronic databases;
12. use Internet databases to gather information on the nucleotide sequence of specified genes, the amino acid sequence of specified proteins, and other molecular information;
13. use alignment software to align DNA sequences downloaded from Internet databases.
Course Content (Lecture):
1. Review of laboratory skills
2. Review of measurements
3. Bacterial culturing techniques
4. Solution preparation
5. DNA restriction digestion and agarose gel electrophoresis of DNA
6. Effects of DNA methylation on restriction enzyme digestion
7. Purification, identification, and quantification of plasmid DNA
8. E. coli DNA
9. Genomic library
10. Restriction mapping of circular and linear DNA
11. PCR primer design using BLAST
12. Extraction and analysis of RNA
Course Content (Laboratory):
1. Micropipetting and sterile techniques
2. Bacterial culture techniques
3. DNA restriction and electrophoresis
4. DNA methylation and restriction
5. Transformation
6. Purification and identification of plasmid
7. pBR322 plasmid restriction mapping
8. Construction of λ genomic library
9. Purification of λ clones
10. Primer design
11. Optimization of PCR for 12S rRNA primers
12. Extraction and analysis of RNA
13. Cell culture
Methods of Presentation:
1. Cooperative laboratory activities
2. Instructor demonstrations
3. Media presentations
4. Lectures and discussions
Assignments and Methods of Evaluating Student Progress:
1. Typical Assignments
a. Reading
1) Textbook
Read chapter 1 and summarize the biotechnology applications used in medical/veterinary and agricultural/food related applications.
2) Articles
Summarize the genesis of the human genome project to the discovery of DNA discussed in this article. Also provide specific examples on how this revelation enabled researchers to understand that human genes are responsible for specific traits.
b. Writing
1) Maintaining laboratory notebook
2) Worksheets, e.g., solving problems and completing study guide reviews
3) Laboratory reports
c. Activities
1) Preparing graphs, tables, and calculations in analysis of laboratory results
2) Measuring mass and volumes to prepare stock solutions and serial dilutions
3) Properly utilizing and maintaining laboratory equipment, e.g., microscopes, spectrophotometer, micropipettors, pH meters
4) Aseptic culturing and staining of microorganisms
5) Accurate laboratory observations
2. Methods of Evaluating Student Progress
a. Examinations, including a final examination, combination of short answer, fill-ins, multiple choice, matching, and essay
b. Professional quality laboratory notebook
c. Quizzes, homework, class participation
Textbooks (typical):
Laboratory DNA Science, Bloom, M., Freyer, G., and Micklos, D., Benjamin/Cummings, 1st edition, 1996*
Basic Laboratory Methods for Biotechnology, Seidman, Lisa A. and Moore, Cynthia J., Prentice Hall, 2nd edition, 2009
* This is the most current edition of the book. It has basic and easy to follow laboratory practices which are currently used in the biotechnology industry.
Special Student Materials:
1. White laboratory coat
2. Safety goggles
3. Disposable latex gloves
4. Bound laboratory notebook
5. Sharpies for marking glassware and Petri dishes
6. Black ink pens for notebook
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