The Biotechnology Curriculum Collection of the California Community Colleges

ORANGE COAST COLLEGE COURSE OUTLINE OF RECORD

Course developer: Farah Sogo
Course static ID:
TOP No. 0430.00
CIP No.
Course adoption:
Date revised:
Revised by:
Semester(s) offered: Fall
Distance education alternative

COURSE MASTER DICTIONARY DATA

Title 5 credit status: Associate degree credit course X
Nondegree credit course
Noncredit course Course name/number: Biology 250
Division: Math and Science
Course title: Laboratory Methods in Biotechnology
Department: Biology
Units: 5
Total course hours: 162
Course length: 16 weeks
Weekly hours configuration: 3.0 hours lecture 6.0 hours lab
Grading method: Graded X
CR/NC
Student option
Noncredit
Method of Instruction: 30 (2 digit no.)
Basic skills status: N (P, B or N)
Materials fee: No
Yes X
$ 12

Justification: X-ray and thermal film, nitrocellulose membranes, and gels are used to analyze DNA and protein, and for permanent reference and technique records.

COURSE PREREQUISITES:
Biology 181, Chemistry 180

CATALOG DESCRIPTION:
This laboratory-based course will be an introduction to modern biotechnological techniques. Labs involve gene cloning and sequencing, restriction analysis, PCR, Southern analysis and genetic manipulation of DNA in bacteria and yeast. Lecture emphasizes the molecular biology that allows these lab applications as well as the molecular biology/genetics of the organisms used.

SCHEDULE DESCRIPTION:
A laboratory-based course which introduces modern biotechnological techniques including gene cloning.

COURSE CLASSIFICATION:

A Liberal arts/AA X
B Remedial
C Remedial
D-H Community course
I Occupational required
I Occupational elective

COURSE TRANSFER:

0 Non-transfer/Non-AA
1 Non-transfer AA
2 Transfer CSU X
3 Transfer UC

JUSTIFICATION FOR THE COURSE:
Required for certification
Occupational state-of-the-art
Advisory committee recommendation

COURSE CONTENT AND SCOPE/TOPIC OUTLINE:
LECTURE OUTLINE

1) Introduction

a) Biotechnology definition
b) Industrial biotechnology
c) Molecular Biotechnology

i) Crops
ii) Drugs
iii) Vaccines
iv) Diagnostics

(1) ELISA

v) Livestock

2) Methods of getting an organism to harbor foreign DNA

a) Transformation

i) Bacteria
ii) Yeast
iii) Transformation Efficiency

b) Plasmids

i) Replication requirements
ii) Physical decryption
iii) Selection requirements and features

(1) In bacteria
(2) In yeast
   (a) Shuttle vectors

3) Genomic Libraries

a) Description
b) How to produce one

4) Gene banks

a) Cloning by complementation

i) Complementing an auxotrophy
ii) Complementing a temperature-sensitive mutant

5) DNA Replication
6) Polymerase Chain Reaction
7) DNA sequences of unknown function

a) Alu family of sequences
b) Variable number of tandem repeats (VNTR's) or minisatellite DNA
c) Satellite DNA
d) Microsatellite DNA
e) Transposons and retrotransposons
f) Pseudogenes
g) Spacer DNA

8) Measuring and Solution making

a) Making solutions of a particular Molarity in a given volume
b) Dilutions from a stock solution
c) How to make percent/volume solutions

9) Transformation into E. coli

a) Calcium chloride
b) Electroporation
c) Conjugation

10) Saccharomyces cerevisiae vectors

a) Cloning vectors
b) Integrating vectors
c) Yeast Artificial Chromosomes (YAC's)

11) Bacteriophage lambda (l) vectors

a) Life cycle of phage l
b) Recombinant phage l
c) Application of recombinant phage l

12) Nucleases

a) Biochemistry
b) Exonucleases
c) Endonucleases

13) Restriction Endonucleases (Type II)

a) Examples
b) Ligation
c) Analysis
d) Restriction mapping

14) E. coli vectors

a) Cloning vectors
b) Cosmids
c) Bacterial Artificial Chromosomes (BAC's)
d) P1-derived Artificial Chromosomes (PAC's)

15) Human Genome Project

a) History
b) Techniques used
c) Current state of

16) Southern Analysis

a) Theory
b) Making probes
c) RFLP Analysis

17) Restriction enzyme analysis of the Globin gene
18) Southern Analysis
19) DNA Subcloning

a) Subcloning the LEU2 gene
b) Blue/White screen
c) Calf Intestinal Alkaline Phosphatase (CIP)
d) T4 DNA Ligase

20) Transfection

a) Establishing cell lines
b) History
c) Cotransfection and selectable markers
d) Transient Gene Expression
e) Role of Mtx and DHFR
f) Episomal Gene expression

21) Transgenic Animals

a) Overview
b) Transgenic mice
c) Knockout Mice
d) Cloning by nuclear transfer
e) Trangenic livestock
f) Trangenic Birds
g) Transgenic Fish

22) Transgenic Plants

a) Purpose
b) Ti Plasmids
c) Using Ti vectors
d) Other plant transformation strategies
e) Industry Applications

23) Dideoxy sequencing

a) Standard Sanger sequencing
b) Cycle Sequencing
c) Automated sequencing
d) Detection of sequencing

24) Primers

a) This lab using Bluescript KS
b) Sequencing a long stretch

i) Making primers
ii) TK and HPRT

c) Making Exos (Nested Deletions)

25) Patents
26) Industry Procedures

a) Standard Operating Procedures (SOP)
b) Good Manufacturing Practices (GMP)
c) Good Laboratory Practices (GLP)

LABORATORY OUTLINE

1) Measurements, Micropipetting, and Sterile Techniques
2) Yeast Transformation of a Yeast Genomic Library
3) Bacterial Culture Techniques
4) Solutions and Dilutions
5) Bacterial mini-plasmid preparations (minipreps) of LEU+URA+ pFL1 genomic library plasmids.
6) Restriction Enzyme analysis of miniprep DNA of LEU+URA+pFL1 genomic library plasmids
7) Southern Analysis of Cleaned up miniprep DNA of LEU+URA+pFL1 genomic library plasmids- Part I

a) Diagnostic digests
b) DNA electrophoresis
c) Electrophoretic transfer to a nylon membrane

8) Southern Analysis of cleaned up miniprep DNA of LEU+URA+pFL1 genomic library plamsids- Part II

a) Probe and Antibody incubations
b) X-ray film exposure and film development

9) Subcloning of a DNA fragment containing a portion of the LEU2 gene in Bluescript KS+
10) Minipreps of Bluescript KS+ ligation with Leu2+containing fragment
11) Restriction analysis of miniprep DNA from Bluescript KS+ ligation with LEU2+ containing fragment
12) Cycle Sequencing the subcloned LEU2+ containing fragment
13) Computer analysis using on-line DNA and protein databases of the sequence.
14) Collaborative Project with Biology 181 students: Quantitation, analysis, and purification of protein and DNA samples prepared by Biology 181 students.

a) Data sheet with sample analysis with pictures will be returned to each Biology 181 lab group
b) Students from Biology 250 will give a short presentation to and field questions from the Biology 181 students.

15) Collaborative Project with Biology 280 students: Biology 280 students prepare Western Blots of globin proteins on nitrocellulose and perform a colorimetric assay. Biology 250 students will reprobe the same blots using a chemiluminescent technique. The results from this analysis will be returned to the Biology 280 students.
16) Collaborative Project with a local high school: The high school students prepare a DNA sample from their hair. The Biology 250 class will perform PCR and gel electrophoresis. The results will be returned to the high school students.

INSTRUCTIONAL OBJECTIVES:

The student will be able to:

1. Describe and perform the fundamental techniques necessary for the purification, analysis and manipulation of DNA.
2. Describe and perform the fundamental techniques necessary to manipulate and analyze protein.
3. Define and apply terminology related to the manipulation of DNA and protein.
4. Describe the methods used to transfer DNA from one organism to another.
5. Apply sterile technique at laboratory sessions.
6. Accurately make and sterilize solutions.
7. Accurately dispense solutions.
8. Practice proper laboratory safety including the proper handling and storage of materials.
9. Define and follow good laboratory practices (GLP), good manufacturing practices (GMP) and standard operating procedures (SOP).
10. Maintain proper laboratory documentation.
11. Analyze DNA and protein sequences using internet databases such as GenBank.

METHOD OF STUDENT EVALUATION:
Student written exams, practical exams, maintenance of a laboratory notebook, presentations to the Biology 181 class, local high school biology class(es), or Biology 280 class, Production of data sheets for Biology 181 and 280, and local high school biology students.

INSTRUCTIONAL METHODOLOGIES:
Lecture on key concepts.
Inquiry based laboratory projects where each lab builds on laboratory products such as plasmids, DNA fragments, as well as bacterial and yeast strains generated in preceding laboratories.
Collaborative projects with other biology classes (Biol 181, 280, local high schools) in which the students send samples to the Biology 250 students for further analysis and data sheet production.
Lab meetings that simulate a research lab meeting and/or a production meeting in industry.
Computer analysis of data using databases available on the internet.

WRITING ASSIGNMENTS/PROFICIENCY DEMONSTRATION:

Students will successfully complete entries on a daily basis into a laboratory book.
Completion of data sheets and written summaries of data provided by collaborative sample analyses.

REPEATABILITY:
N/A

FEASIBILITY:

Faculty: This course will be taught using existing faculty.
Classroom: This course will be taught in existing classroom facilities
Library Learning Resources: The library has an interlibrary loan program and internet access that will be
sufficient for any necessary classroom research.

EDUCATIONAL MATERIALS:
Molecular Biotechnology: Principles and Applications of Recombinant DNA, Glick, B.R. and Pasternak, J.J., ASM Press, Washington, D.C.
Molecular Biotechnology: A lab course for undergraduates using bakers yeast as a model system, Sogo, L.F.