How to Start a Biotechnology Program

Mary Jane Kurtz, Ph.D NBC 2 Consultant [email protected]

Why Biotechnology?

A number of good reasons

1. Academics 2. Technical skills 3. Biotechnology as a future career 4. Biotechnology is exciting Biology can be seen with a molecular perspective Can answer questions about ourselves Can provide new medical aid to the afflicted

Getting Support from Your Administration

This is important to the future success of your new program Allows for obtaining outreach to new students, space to grow, and expenses for needed equipment Supports the teachers who are involved in program

Academic Advantages

 Allows students to do up-to-date hands-on experiments  Demonstrates the integration of all sciences, math, and engineering through experiences in the laboratory  It develops writing skills for clarity, focus and documentation

Technical Advantages

 Curriculum can introduce career tracks for students in a developing scientific area  Using hands-on activities, students have a opportunity to develop skills needed in the workplace at all levels of formal training  Potential career paths flowing from the knowledge gained is broad based

Biomanufacturing Curriculum: National Standard Concepts Covered

• Math, Biology, Chemistry in Curriculum – Measurement – Solutions – Enzyme reactions – Transformation of cells with DNA – Forces used in centrifugation, electrophoresis etc.

• National Academy of Sciences Standards – Unit of Math & Science – Structure/properties of matter – Chemical reactions/conservation of matter – Cell structure and functions, heredity – Motions and forces

Biotechnology A plus for Teachers and Students

   Integrated science education: Science Technology Engineering and Math (STEM) Career pathways towards work/school focused on science with thousands of new jobs predicted in the next few years Laboratory based activities = increased interest – More exciting ways of introducing concepts – Hands-on learning is more inclusive – State-of-the-art laboratories

Biomanufacturing vs Biotechnology

     Offers diverse career pathways Hands-on learning of science Understanding of good laboratory practices Stability in career pathways Assistance in obtaining college degrees through company support  Both highly academic and technically oriented students

Ten Technician Jobs Anchor Ten Biomanufacturing Departments

• • • • • • • • • Facilities/Metrology Validation Environmental Health and Safety (EH&S) QA

Upstream Processing Downstream Processing QC Microbiology QC Biochemistry

Process Development

RESEARCH & DEVELOPMENT(pre-clinical):

Discovery Research

OPERATIONS:

Process development, Manufacturing & Production

QUALITY:

Quality Control & Assurance

CLINICAL RESEARCH:

Clinical Research: Regulatory Affairs

Discovery Research

Senior Scientist Scientist III,II,I Research Associate

Bioinformatics

Scientist Engineer Analyst Programmer

Process Development

Director Supervisor & Process Development Technician

Manufacturing & Production

Supervisor Associate Technician (Operator) Instrumentation Tech Calibration Technician

Facilities Management

Manager Facilities Technician Shipper/receiver • Careers in red indicate entry level positions All above require 2-4 year degrees Entry level positions require an Assoc deg or certificate

Quality Control (QC) Clinical Research

Clinical Research Manager

Chemistry

Chemistry QC Analyst QC technician

Microbiology

Microbiology QC analyst QC Technician

Quality Assurance (QA)

Documentation Specialist QA Documentation Coordinator

Clinical Research

Clinical Research Manager

Regulatory Affairs

Manager Associate Data Manager

Business Development

Director of Business Development

Administration

Human resources Safety Manager Higher entry levels require a BS, MS, PhD or Engineering degrees Can require medical, nursing or business degrees

Student Success Stories

Students completing courses with certificate or Associate degrees in Biomanufacturing: www.biomanufacturing.org

http://www.biomanufacturing.org/

1. How to Start A Biotechnology Program

(After getting support from the administration) Gain the support the Industry in your Area: – Find out the location and focus of each of the biotechnology/support companies within (10) miles of your school – Understand the desired skills individuals (such as your students) should have (Most important of these are soft skills) – Organize an advisory group that can assist you in curriculum building towards skills needed This will be a source of information as well as help with technical supplies

2. Find or Develop a Curriculum

Resources

High School Level-Entry Level Community College: Protein is Cash: Northeast Biotechnology Center and Collaborative , NBCC: www.biomanonline.org, Biomanufacturing.org

Biotechnology: Science for the New Millennium : Ellyn Dougherty, EMC publisher, www.emcschool.com Basic Laboratory Methods in Biotechnology:Textbook and Laboratory Reference: Lisa Seidman et al, Prentice Hall, ISBN 013-795535-9

Align Curriculum with Program Goals

 What are your goals?

– Two year associate degrees transfer to four year college – Two year associate degree and work – Certificate and immediate work  Research and Development or Manufacturing?

 Goals can vary but should allow majority of your students to participate in.

Four year college career pathway Biology Undergraduate Other Undergraduate or Engineer Biomedical Ph.D student Other Ph.D student Industry Junior Postdoc & Senior Postdoc Other Science Position Staff Scientist Assistant Professor Independent Investigator

Intro to Biotechnology for High School/CC

1 2 3 4 10 11 5 6 7 8 9 12 Introduction to Biotechnology Metrology Basic Microbiology Unit 1 Basic Microbiology Identification of Bacterial Types Unit 2 Transformation of Cells With DNA Spectrophotometry Genetic Transformation of Bacteria The Rain Forest:How do we find new pharmaceuticals ?

Proteins and their purification Good Manufacturing Practices Documentation Chromatographic separation and how it works QC Sampling of Product to Determine its Purity Use of bacteria, enzymes and fermentation by yeast Validation of Pipets and Calibration SOP ‘s & Documentation of Testing Basic Aseptic Technique and Testing of Bioload in Biomanufacturing Areas Gowning up Identify of Microbe Morphology and Electrophoresis PGlO transformation of Bacteria Protein determination of samples By Bradford and UV analysis E. coli transformation with pGLO plasmid Growth of pGlo transformed bacteria and isolation of the p-GLO product Isolation of casein from milk Using solubility and filtration techniques GMP activity with popcorn And use of Documentation Use of ion exchange and hydrophobic columns in Isolating GFP Polyacrylamide Gel Electrophoresis

Biotechnology Curriculum

2. Associate Degree or Certificate Program Resources: Introduction to Biomanufacturing: Global Biomanufacturing Lab manual NBC2, at www.biomanufacturing.org

Introduction to Biomanufacturing: Textbook, NBC2, at www.biomanufacturing.org

Introduction to Biotechnology Lab Manual ATE central: Linnea Fletcher, et al ISBN BITC1311006

2 3 4 5 6

Techniques in Biotechnology

1 Acids, Bases and Buffers Protein & Spectrophotometry Enzyme Kinetics Kinetics of LDH Purification by pH Column Chromatography Enzyme Purification by Ion Exchange Identification of Proteins High Pressure Chromatography Preparation of Buffers Determination of Protein in solution Protein structure Km of enzymes Precipitation of pro tein and Affinity chromatography Purification of LDH using a IEX column Concentration and Electrophoretic Techniques Quality control

Case study of Two Different Programs

• • • •

College Degree

Requirements will be more demanding in math, science Comprehension, graphing &computer work is good can work independently Has appropriate soft skills and can work well as a team member Is good at trouble shooting problems in labs • • • • •

High School or Certificate

Entry level math is pre algebra or algebra 1 Science background is sketchy (lacks chemistry) Desire to be placed into a position after graduation Likes science but needs reassurance Needs to have SOP to carry out work

High School Curriculum

A Workshop to Instruct Teachers and Students in Biomanufacturing and the Bioeconomy

Career Tracks: Protein is Cash

Introduces

     Upstream Processing – Production of pGLO into protein by transformed cell Downstream Processing – Separation of cellular debris and cell supernatant – Purification of pGLO protein by Chromatography Quality Control – Identification of protein product by electrophoresis Discovery Research – How new drugs are discovered Biofuels

1. Metrology 2. Transformation of Bacteria 3. Upstream Processing 4. Downstream Processing 5. Quantitative Analysis 6. Discovery Research

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Protein is Cash

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Local Teachers Workshops

Biomanonline.org

Day 1 : Metrology/Instrumentation

Activities

• Calibration of top balance • Verification of pipet performance • Pipetman Challenge

Outcomes

• Introduction of Good Manu facturing Practices • SOP and documentation • Confidence in measurements by instru mentation

Metrology

Day 2 Transformation of Bacteria

• • • • •

Activities

Addition of pGLO plasmid to bacteria in Ca++ solution Heat /shock the mixture Plate cells onto selective Luria broth agar + ampicillin Incubate overnight at 37oC Note: arabinose acts to turn on the gene which starts production of pGLO protein • • • •

Outcomes

Selection of cells by growing on ampicillin plates Only transformed with cells will survive due to amp-r gene Selected colonies will multiply in Luria broth at 37oC overnight Aseptic technique and proper disposal of bacteria

Day 2 E.coli Transformation with pGLO plasmid

Ampicillin resistance gene (Amp r ) and target gene on bacterial plasmid Bacterial clones Cell division Only E. coli containing plasmid survive on Ampicillin plates Transformation mixture is plated on to agar plate containing Ampicillin

Results of Inserting Foreign DNA into an Organism

Cells will multiply and produce desired gene product pGlO gene expression vector: Green Fluorescent Protein

Day 2: Upstream Processing: Cell Growth and Lysis

• • • •

Activities

Transformed cells grown overnight in selected media are separated from media by centrifugation Media is removed and packed cells are lysed Homogenate is centrifuged Supernatant with pGLO protein is retained for downstream processing • •

Outcomes

Multiplication of cells Initial separation of fluorescent protein from cell homogenate

Day 3: Downstream Processing:

Purification of Green Fluorescent Protein by Chromatography •

Activities

Separation of product by Different types of Chromatography •

Outcomes

Fractions with green fluorescent protein will glow and be selected for analysis – – – – Size exclusion Hydrophobic Interaction Cationic Interaction Anionic Interaction • Understand concept of chromatography as selective interactions of compounds with matrix

Isolation and Purification of Green Fluorescent Protein

Transformed cells Test tubes Fraction number #1 #2 #3 Courtesy of Bio-Rad

Separation of Proteins by Column Chromatograph

 Separation by size – Size exclusion chromatography  Separation by hydrophobic characteristics – Water loving vs water hating environments  Separation by ionic charge – Protein has a positive charge & attaches to negative matrix on column: Cation Exchange – Protein has a negative charge & attaches to positive matrix on column: Anion Exchange

Size Exclusion Chromatography

Red molecule =10^6 daltons - Blue molecule = 600 daltons

How Size Exclusion Works

 Molecular size of molecule will separate two or more molecules  Large molecules can not go into a bead of a certain size and flows quickly through a column  Small molecules enter into a bead and flows slowing through a column.

 Size of two different molecules are separated

H+ H+

Hydrophobic Interaction Chromatography

H+ H+ Hi Salt Low salt High salt: 2 M (NH 4 ) 2 SO 4 Wash buffer: 1.3 M (NH 4 ) 2 SO 4 Elution buffer: 10 mM Tris

Ion Exchange Chromatography

• • • Proteins bind to opposite charges on the matrix An example of ion Exchange Addition of increasing Salt/pH should release proteins

Cation Exchange vs Anion exchange

 How proteins are attracted to ion exchangers

Day 4: Quality Control

• • • •

Activities

Chromatography fractions prepared for electrophoresis Electrophoresis Box is assembled with PAGE gel Samples of chromatography fractions are added to PAGE gel and ran for 30 minutes Gels stained and viewed • • •

Outcomes

Analysis of protein samples by observation on light box determines protein purity Standard molecular weight markers indicate size of protein Verification of mol wt by comparison with standard proteins and number of proteins in a single sample

Quality Control Analysis of Column Fractions

 Isolated fractions using Ion Exchange Chromatography are then analyzed  Electrophoresis by SDS PAGE of fractions collected

Day 5. Discovery Research and FDA approval process

• • • •

Questions

Discovery Science in Drug development Good Manufacturing Practices and the FDA Initial Biomanufacturing Process Development Scale up to full production of biologic /clinical trials • • •

Answers

Visit local biomanufacturing plants Discuss importance of documentation in the workplace ie. SOP and Batch Records Invite industry representatives to speak to the regulated workplace

Bio-Rad Partnership

Northeast Biomanufacturing Center and Collaborative (NBC 2 ) is partnered with Bio-Rad:  Provides teachers with engaging hands-on  biomanufacturing education that is easily accessible Introduction to: – – – metrology, production (upstream and downstream processing) quality control biochemistry and clinical trials. These modules can be brought directly into the classroom

Virtual Chromatography http://www.Atelearning.com/BioChrom

HTTP

Useful Information for Biomanufacturing

   http://www.Biomanufacturing.org

http://www.Biomanonline.org

http://www.Bio-link.org

Background Resources: “Development of Biotechnology Curriculum for the Biomanufacturing Industry”, Robert McKown, and George L. Coffman, May/June 2002, Pharmaceutical Engineering pages 1-6.

“ Introduction to Biomanufacturing a Global Biomanufacturing Curriculum” Northeast Biomanufacturing Center & Collaborative (NBC2) 2011, publisher :Lulu.com