BIOTECHNOLOGY

Learning Goals Describe the science of Biotechnology and its

product domains

List the steps to producing a GMO through use of rDNA

Outline the steps of producing and delivering a product made through recombinant DNA technology

Identify the important labs and agencies in the US, and what they’re mainly responsible for

Learning Goals cont.

Apply the strategy for values clarification to bioethical issues

Identify products made through biotechnology

Identify products made through biotechnology

Vocabulary Insulin Biotechnology DNA rDNA technology PCR Cloning Fermentation Diabetes Antibiotics Restriction enzyme DNA ligase rDNA

Proteases Antibodies Pharmaceutical R&D Pure Science NIH CDC DNA Fingerprinting Microbial agents Virus Applied science

Vocab cont. Moral GMO E. coli Plasmid t-PA Efficacy Clinical trials FDA Cystic fibrosis Biochemistry Molecular biology Genetics ethics

Therapeutic EPA USDA Data Hypothesis Variable Control Positive control Negative control Concentration Journals Genome bioethics

Defining Biotechnology

We have been manipulating entire organisms for thousands of years through selective breeding to produce plant and animal products that are Bigger Healthier More productive Have novel phenotypes

Scientists recently learned to manipulate not only whole organisms, but also the molecules, cells, tissues, and organs of which they are built

Example: Human Insulin

Insulin is a protein produced in healthy individuals that facilitates the uptake of sugar from the blood stream into specific tissues

In some diabetic patients, the ability to produce insulin is impaired

In the past diabetic patients had to take injections of Bovine (cow) insulin to regulate their blood sugar

Example: Human Insulin

Problem: getting insulin from cows is not very efficient, it’s time consuming and expensive, and can lead to improper immune stimulation of the recipients

Solution: Biotechnology We have learned to manufacture large

amounts of specific molecules, such as human insulin in bacterial cells Cost effective Convenient Fast

Biotechnology: Any technological application that uses biological systems, living organisms, or derivatives thereof, to make or modify products or processes for specific use

PAST PRESENT

Selective breeding Fermentation: a

process by which in an oxygen deprived environment a cell converts sugar into lactic acid or ethyl alcohol (ethanol)

DNA technology Recombinant DNA

(rDNA) technology PCR Cloning Proteomics

Modern Biotechnology

Since biotechnology is constantly changing there is no one definitive definition

In 2003 in an international treaty on biotechnology, modern biotechnology was defined as : “the application of techniques that overcome

natural physiological reproductive or recombination barriers and that are not techniques used in traditional breeding and selection “ including In vitro nucleic acid techniques The fusion of cells beyond taxonomic family

Restriction Enzymes

One major discovery that propelled the biotechnology science and industry was the discovery of restriction enzymes

Restriction enzymes: enzymes that cut DNA at specific recognition nucleotide sequences known as restriction sites

They were discovered in bacteria & are thought to have evolved to provide a defense mechanism against invading viruses

Recombinant DNA Technology

Restriction enzymes allow us to cut and recombine DNA molecules, often from different species

“cutting” Reminder DNA (deoxyribonucleic acid) is a double

stranded helical molecule that that stores genetic information for the production of all of an organisms proteins

DNA Ligase: an enzyme that binds the disconnected strands of DNA together “pasting”

If you put DNA into an organisms cell, that cell will produce the protein that DNA codes for!

Importance of rDNA technology

Advances in rDNA technology allow biotechnologists to manufacture a wide variety of products either previously unavailable or that could only otherwise be made in small quantities

Examples: Insulin: for treatment of diabetes Proteases: function in breaking down other

proteins Antibodies: proteins normally developed by the

immune system for recognizing specific molecules called antigens on infectious agents and so limiting their infectious ability

Other Helpful Techniques

PCR: Polymerase Chain Reaction is a technique that involves copying short pieces of DNA and using them as templates to make millions of copies in a very short amount of time

Cloning: method of asexual reproduction that produces identical organisms Done mainly in bacteria and some fungi Only happens in animals if manipulated by

scientists to do so

Cao Yilin, a plastic surgery expert in Shanghai, successfully reproduces a human ear on the body of a little white mouse on Friday, April 4, 1997 in Shanghai.

Dolly, right, the first cloned sheep produced through nuclear transfer from differentiated adult sheep cells, and Polly, the world's first transgenic lamb, are in their pen at the Roslin Institute in Edinburgh, Scotland, in early December, 1997.

Biotechnology Workplace

Biotechnology is practiced in a variety of settings , and in general the setting determine the major emphasis Private companies: focus is making money so

they tend to emphasize the development and manufacture of products that will yield high profit margins The do have a Research & Development department,

but they will only fund research that has potential for financial pay off

Universities & Government agencies focus on research More involved in Pure science

The main difference between companies and university or

government labs is that companies must provide a

product or service that results in earnings, a nonprofit research

facility does not

Pure vs. Applied Science

Pure-science: Scientific research whose main purpose is to enrich the scientific knowledge base

This is the type of science nonprofit labs focus on

Applied Science: the practice of utilizing scientific knowledge for practical purposes, including the manufacture of a product This is the type of research forprofit labs focus

on

4 Main Categories of Biotechnology Companies

Pharmaceutical Agricultural Industrial Research or production instruments,

reagents, or data Some companies sell their services rather

than product Example: 23 and Me See figure 1.13

R&D vs. QC

Research & Development : refers to the early stages of product development that includes discovery of the structure and function of a potential product and the initial small scale production

Quality Control is a measure of precision or how well the measurement system reproduces the same result over time and under varying operating conditions

Important Government Labs

National Institute of Health (NIH): the federal agency that funds and conducts biomedical research

Centers for Disease Control & Prevention (CDC): National research center for developing and applying disease prevention control, environmental health, and health promotion and educationactivities to promote public health

Bioengineered Products

By the 1970’s scientists had developed new methods including the use of restriction enzymes and DNA ligase to create new combinations of genetic information, or rDNA

rDNA can be inserted into cells giving these cells a new genotype phenotype

Organisms containing these modified cells are called Genetically Modified Organisms (GMOs)

GMOs

GMO’s contain DNA from another organism and will produce the new proteins encoded in the acquired DNA

The 1st GMO’s to produce a human protein were Escherichia coli (E. coli) bacteria

E. coli bacteria are the work horse of molecular biology, no other bacteria is used as frequently as this one

Escherichia coli

Were given pieces of human DNA (genes) containing instructions to produce a human growth hormone called somatostatin

The somatostatin gene was carried into the E. coli cells on tiny pieces of bacterial DNA called Plasmids These recombinant DNA plasmids contained

both bacterial and human DNA The E. coli cells read the human DNA and

produced the human protein somatostatin, which could then be harvested and purified for use

Tissue Plasminogen Activator

One of the 1st genetically engineered products to be sold was t-PA

t-PA is a blood clot dissolving enzyme that can be used immediately after a heart attack to clear blocked blood vessels The body produces only small amounts of t-PA To produce enough t-PA for therapeutic use

scientists genetically engineered mammalian cells using Chinese Hamster Ovary (CHO) cells

Chinese Hamster Ovary

Ovary cells are grown in culture and engineered with the gene that provides the instructions to make the t-PA enzyme

By subjecting the cells to the right conditions, the CHO cells accept and incorporate the foreign DNA and begin the process of transcription and translation

Large amounts of t-PA can then be extracted and purified See figure 1.18

Biotechnology & Society

Applications of rDNA and genetic engineering have resulted in the creation of hundreds of Biotech companies , specializing in all kinds of GMOs and their protein products including Proteins used in pregnancy tests Enzymes that increase the amount of juice that

can be extracted from an apple Molecules used in vaccines

In 2004 there were already over 370 drug products created from rDNA techniques undergoing clinical trials

Strawberry plants that can grow in freezing weather

Biotechnology & Healthcare

Biotechnological advances will enable patients to submit tissue samples and receive a treatment regimen designed specifically for them. This is calle Personalized medicine, or pharmacogenomics pharmacogenomics: pharmaceuticals based

on genomics Present research based on broad

characteristics such as ethnicity, gender, and family history has already been performed and treatments have been modofied accordingly

The Human Genome Project

The human genome project was an international effort to sequence and map all the DNA on the 23 pairs of human chromosomes

We know the sequence of the human genome, but we still do not know all of the sequences that code for proteins For the ones we do know about, we don’t

completely understand how, when, and where they are turned on or off

This is called differential gene expression DNA intro

Biotechnology & Healthcare

As sequencing human genomes becomes more cost effective, there will come a time when drug regimen is determined by the genotype of the patient or the tumor being targeted

One of the 1st drugs to accomplish this is the drug Herceptin which will only target breast cancer cells that produce too much of the protein HER2

The decision to administer Herceptin as part of the chemotherapy regimen is based on a single gene

Biotechnology & Healthcare

Some drugs, like chemotherapeutic agents, have serious side effects because they treat the whole organism and not just the diseased tissue

Advances in biotechnology have enabled the development of therapies that can be delivered directly to diseased tissue and greatly reduce side effects Rexin-G: a drug that targets pancreatic cancer is

delivered by a genetically engineered viral particle that is designed to seek out and kill metastasized cancer cells is in clinical trials now

How Companies Select Products for Manufacture

Research & Development No matter what the product, the goal is

to make it and market it as quickly as possible

The R&D phase for a new pharmaceutical often takes from 10-15 years for a company to move a product through the “pipeline”

Product Pipeline

A drug must demonstrate “proof of concept” data in the research phase before being allowed to move to the development phase, including Is it feasible to manufacture in amounts

sufficient to treat people What needs to be done to ensure safety What characteristics indicate it will be

effective And unfortunately, what is the profit margin This is accomplished with a Product

development Plan

Product Development Plan

Does the product meet the following criteria Does the product meet a critical need? Is there a market large and wealthy enough to

produce enough sales Does preliminary data support that the product will

work? Can patent protection be secured Can the company make a profit?

Each product in a pipeline will be reviewed regularly and dropped from production if the answers to these questions are unsatisfactory

See figure 1.21

Product Pipeline

Projects then enter clinical development Testing is done small scale before large scale

production is pursued All pharmaceuticals have to undergo at least 3

rounds of strict clinical trials under the guidance of the FDA before being marketed

Example: Pulmozyme is manufactured by Genentech Inc and is an enzyme used to treat patients with Cystic Fibrosis People with CF experience a build up of mucus in

the lungs and intestines and is often fatal by the age 30

Pulmozyme reduces the amount of mucus produced

In 2003 Pulmozyme had

sales of $167million

Regulations Governing Product Development

All biotechnology products have regulations governing their production in the product pipeline Environmental Protection Agency (EPA)

The federal agency that enforces environmental laws including the production and use of microorganisms, herbicides, pesticides, and GMOs

United States Department of Agriculture (USDA) Federal agency that regulates the use and

production of plants, plant products, plant pests, veterinary supplies and medications, and genetically modified plants and animals

Biotechnology with a Conscience

Being able to distinguish between right and wrong and to make decisions based on that knowledge is considered having good morals

Problem: not all people have the same list of right and wrong, and so we often have different morals

Example: Vegans don’t eat or use any animal products, and many consider it immoral to do so

Fact: most people eat and use animal products on a daily basis without any qualms

Also Fact: those same people that

don’t have any problem eating meat and using animal products

would be horrified to think of going

home and cooking their pet dog

Sparky or their new kitten Mittens for

dinner

Bioethics

The study of moral standards and how they effect conduct is called ethics

Bioethics is a term that has been coined to describe the study of decision making as it applies to moral decisions that need to be made because of advances in biology, medicine, and technology

Bioethics

Many of the new biotechnologies are controversial because they force people to think about what they believe is right or wrong

Examples: Harvesting and manipulating embryonic stem

cells It kills the human embryo

Genetically modifying food Are they really safe to eat

Prenatal genetic testing/ engineering Playing God

Technology is Not Evil

New technologies generate ethical questions

The positions one takes on ethical issues are based on how you feel about the knowledge and understanding you have of a subject

There is often not a clear right or wrong answer

Most of the time you have to make a choice between the lesser of 2 evils

So how do we decide what’s “right”

But sometimes people are!

Strategy for Values Clarification

Identify and understand the problem or issue. List all the possible solutions to the issue Identify the pros and cons of each of the

solutions Consider legal, financial, medical, personal, social,

and environmental aspects Based on the pros and cons for each solution,

rank all the solutions from best to worst Decide if the problem is important enough to

take a position. If it is, decide your position and be prepared to describe and defend it

Doing Biotechnology: Scientific Methodology in a

Research Facility There is no 1 correct scientific method The scientific method is really just a

series of common sense steps and techniques that scientists use to solve a problem or answer a question

There are several practices used by most scientists when conducting experimental research

Scientific Methodology

State a testable scientific question or problem based on some information or observation

Develop a testable hypothesis Plan a valid experiment

Multiple replicates Control groups

Conduct the outlined experiment and collect and organize data

Formulate conclusions based on experimental data and error analysis

Control Groups

POSITIVE CONTROLS NEGATIVE CONTROLS

A group that will give a predictable positive result

A group that will give a predictable negative result

Writing Experimental Conclusions

A good approach to writing an experimental conclusion is to use the REE/PE/PA method

REE: Results with Evidence and Explanation PE: Possible Errors

Identify the sources of expreimental design errors that would or could lead to false or misleading data, and explain possible implications from such errors

PA: Practical Applications: discuss the meaning or value of experimental results in the

short and long term. Identify how the findings could be valuable clinically, to the

company, or to the community Identify the direction the experiment will go take from

here

Review: Describe the science of Biotechnology and its product domains

Biotechnology: The study and manipulation of living things or their component molecules, cells, tissues, or organs

Product Domains Pharmaceutical Agricultural Industrial Research or production instruments, reagents,

or data

Review: List the steps to producing a GMO through use of rDNA

Identify a gene of interest Treat gene of interest and plasmid

vectors with a restriction enzyme Use DNA ligase to glue gene of interest to

plasmid Insert plasmid into host organism (plant,

animal, or bacteria) through various mechanisms

The rDNA will be used by the host organism to produce the new protein of interest

Review: Outline the steps of producing and delivering a product made through recombinant

DNA technology

Come up with an Idea Research and Development

Proof of Concept through a Product Development Plan Will it work Is there a market for it Can we make it in sufficient quantities

Large Scale Production Clinical Trials FDA approval

Review: Identify the important labs and agencies in the US, and what they’re mainly

responsible for

NIH: the federal agency that funds and conducts biomedical research

CDC: National research center for developing and applying disease prevention control, environmental health, and health promotion and education activities to promote public health

EPA: The federal agency that enforces environmental laws including the production and use of microorganisms, herbicides, pesticides, and GMOs

USDA: Federal agency that regulates the use and production of plants, plant products, plant pests, veterinary supplies and medications, and genetically modified plants and animals

Review: Apply the strategy for values clarification to bioethical issues

Identify /understand the issue List all the possible solutions Identify the pros and cons of each considering

legal financial, Medica personal Social environmental aspects

Rank solutions from best to worst Decide and defend your position

Identify products made through biotechnology

T-PA Human insulin Strawberry plants that can grow in

freezing weather Pulmozyme