BOT106 - Protein-Based Technology 2

Outline info
Last revision date 2018-07-20 11:36:18.948
Last review date 2018-07-20 11:36:26.374

Subject Title
Protein-Based Technology 2

Subject Description
In this course students learn about the production and expression of recombinant proteins. Topic include; gene amplification and sub-cloning into expression vectors, expression and purification by His-Ni affinity chromatography; qualitative and semi-quantitative analysis by Western blot, ELISA and radio-immunoassays (RIA); examples of applications in different biotechnology sectors; laboratory practices.

Credit Status

Learning Outcomes
Upon successful completion of this subject the student will be able to:

  1. Examine the production process of recombinant proteins
  2. Explain the theoretical and practical principles of gene isolation and sub cloning strategies into expression vectors.
  3. Perform different methods of protein purification using affinity chromatography.
  4. Design and assemble DNA expression vectors.
  5. Produce recombinant proteins using different expression vectors.
  6. Identify and quantify recombinant protein products

Cheating and Plagiarism
Each student should be aware of the College's policy regarding Cheating and Plagiarism. Seneca's Academic Policy will be strictly enforced.

To support academic honesty at Seneca College, all work submitted by students may be reviewed for authenticity and originality, utilizing software tools and third party services. Please visit the Academic Honesty site on for further information regarding cheating and plagiarism policies and procedures.

All students and employees have the right to study and work in an environment that is free from discrimination and/or harassment. Language or activities that defeat this objective violate the College Policy on Discrimination/Harassment and shall not be tolerated. Information and assistance are available from the Student Conduct Office at

Accommodation for Students with Disabilities
The College will provide reasonable accommodation to students with disabilities in order to promote academic success. If you require accommodation, contact the Counselling and Disabilities Services Office at ext. 22900 to initiate the process for documenting, assessing and implementing your individual accommodation needs.

Completion of BOT 100

Topic Outline
Week 1: test and assignment dates. Applications of protein engineering in biotechnology, examples in pharmaceutical and food industries; assignment of related topics for students’ presentations. Laboratory: selection of expression vectors and restriction enzymes for gene sub-cloning; vector and insert restriction, isolation and purification using preparative agarose gels.

Week 2: practical protocols for production of recombinant proteins, sub-cloning strategies; elements for expression: bacterial, yeast, vaculovirus, plant and mammalian vectors; commercially available reagents and kits. Laboratory; sub-cloning of restricted DNA and transformation of competent cells.

Week 3: genetic transformation, methods and mechanisms; natural and artificial competence; preparation of competent cells. Laboratory: selection, isolation and analysis of clones containing a recombinant protein by SDS-PAGE.

Week 4: PCR amplification theory (review); sub-cloning of PCR products into expression vectors; construct and primers design. Laboratory: PCR amplification of a DNA fragment encoding the firefly luciferase gene and sub-cloning into an expression vector.

Week 5: transformation of competent cells with the firefly luciferase gene; midterm test-theory and practice.

Week 6: review of protocols for PCR screening of transformed bacterial colonies. Laboratory: selection and identification of positive clones by PCR and confirmation of gene expression.

Week 7: biosynthesis of recombinant proteins containing peptide tags; principle and methods; review of different peptide tags, their use and applications for detection, solubilization, and affinity chromatography purification of recombinant proteins. Laboratory: affinity purification of a His-tagged and a GST-tagged recombinant protein by affinity chromatography. 

Week 8: principles of immunodetection; Radioimmune Assay, Enyzme Linked Immune Sorbant Assay (ELISA) and Western blot: Laboratory: detection of affinity chromatography-purified recombinant proteins by Western blot.

Week 9: completion of previous laboratory exercise and review of experimental results; students’ presentations.

Week 10: final exam-theory and practice.

Mode of Instruction
Short lectures, laboratory practices, tutorials and assignment presentations. Discussions

Prescribed Texts
To be introduced by the instructor

Reference Material
Prescribed text books, Laboratory experiment notes in form of procedures and reports

Required Supplies
Laboratory coat, safety glasses

Promotion Policy

Grading Policy
A+ 90%  to  100%
A 80%  to  89%
B+ 75%  to  79%
B 70%  to  74%
C+ 65%  to  69%
C 60%  to  64%
D+ 55%  to  59%
D 50%  to  54%
F 0%    to  49% (Not a Pass)
EXC Excellent
SAT Satisfactory
UNSAT Unsatisfactory

For further information, see a copy of the Academic Policy, available online ( or at Seneca's Registrar's Offices.

Modes of Evaluation

  • Assignments are due at the beginning of the class on which they are due.
  • A late penalty of 10% per day is assessed for late assignments, including those not handed in at the beginning of class when due.
  • Material will not be accepted after one week following the due date and/or when the marked material is returned to students, whichever comes first.
  • Assignments are to be prepared by computer.

Absenteeism and Exams/Quizzes
  • Students should be aware that absenteeism almost guarantees an inability to achieve satisfactory grades.
  • Students who are absent for an examination due to an emergency (e.g., motor vehicle accident, hospitalization or death in the family) may provide official documentation within five days of the missed exam and be provided a deferred exam at a later date.  Official documentation includes a death notice or an original doctor’s certificate identifying the date, length of time expected absence and the specific reason for the absence.  Examinations missed without official documentation and approval result in a grade of zero.
  • There are no deferred options for missed quizzes.  The value of missed quizzes, at the discretion of the Faculty, will be added to other evaluation components.

English Proficiency
  • All written work should demonstrate the following characteristics for clarity and conciseness:
-writing is consistent with the rules of English grammar
-spelling and punctuation are correct
-sentences are structured correctly
-main ideas are supported with specific, relevant examples and reasons
-work flows logically through supporting statements/paragraphs
-work is arranged in correct format (e.g., as a report, essay)
-up to 10% of the final grade may be deducted on all work if the above English competencies are not met.

Format for Assignments
  • Students must use the standard, APA style for quoting sources.   Help is available at:

Laboratory Attendance
The laboratory component is essential and therefore it is strongly recommended that you attend all labs.  Any missed labs must be supported with a legal document with three days of the lab.  Any student who fails to attend more than 2 laboratory classes for a 10 week subject will not pass the subject.    

Laboratory Safety
Students are required to review and understand the safety procedures and guidelines outlined on the first class and sign the sheet to this effect before beginning work in the laboratory.  Students must also wear a lab coat and safety glasses when conducting experiments.
A student who arrives without the proper safety equipment will not be permitted to participant in the lab but will be asked to leave the class.  The student will receive no grade for the lab missed.

Grading is based on the following marking scheme:
Laboratory reports 20%
Assignments   10%
Midterm test 30%
Final Exam  40%

Note: Two Labs (experimental reports) each worth 20% of final mark. The requirements of the reports are presented in writing at the beginning of the class.


Other Evaluation Considerations
The student is expected to comply with the Safety Rules for working in the laboratory, sign the safety contract, and WILL NOT UNDER ANY CIRCUMSTANCES WEAR CONTACT LENSES in the laboratory. The student will know where all safety equipment is located in the laboratory and will be familiar with WHIMIS concepts and signage.


Approved by: Denis Gravelle