Today’ s Agenda: Journal Questions:

(1) Describe what you know about DNA.

(2) What is genetic engineering (Biotechnology)?

*1. Lecture: Genetic Engineering (Biotechnology) & Recombinant DNA Technology -slide …. 80

04/21/23 1

Genetically Modified (GM) Crops around the World

04/21/23 2

04/21/23 3

04/21/23 4

04/21/23 5

Genetic Engineering

Dr. Rick Woodward

Genetic Engineering

04/21/23 6

Genetic Engineering

04/21/23 7

Genetically Engineered Boneless Chicken Ranch

04/21/23 8

DNA, the Law, and Many Other Applications – The Technology of DNA Fingerprinting

04/21/23 9

A DNA fingerprint used in a murder case.

The defendant stated that the blood on his clothing was not his.What are we looking at? How was it produced?

DNA Fingerprinting Basics

04/21/23 10

A. Different individuals carry different alleles.

B. Most alleles useful for DNA fingerprinting differ on the basis of the number of repetitive DNA sequences they contain.

DNA Fingerprinting Basics

04/21/23 11

A DNA fingerprint is made by analyzing the sizes of DNA fragments produced from a number of different sites in the genome that vary in length.

The DNA Fragments Are Separated on the Basis of Size

04/21/23 12

The technique is gel electrophoresis. The pattern of DNA bands is compared between each sample loaded on the gel.

Gel ElectophoresisA. Technique used to separate

nucleic acids or proteins by size and charge.

04/21/23 13

California Biology Content Standards:

California Content Standards:

5 b. Students know how to apply base-pairing rules to explain precise copying of DNA.

5 c. Students know how genetic engineering (biotechnology) is used to produce novel biomedical and agricultural products.

5 e. Students know how exogenous DNA can be inserted into bacterial cells to alter their genetic makeup and support expression of new protein products.

04/21/23 14

DNA Review:A. Structure: Double Helix

B. Location: Nucleus of Cell

C. Function: “Blue print of Life” – Creation of Proteins/Amino Acids – Transcription.

D. Nitrogen Base: ATCG

E. Nucleotide: Phosphate, Sugar (Deoxyribose), Nitrogen Base (ATCG).

F. Base Pairing Rules: A-T

C-G

04/21/23 15

Genetically Modified (GM) Food

04/21/23 16

Genetically Modified Cotton (contains a bacterial gene for pest resistance)

Standard Cotton

Genetically Modified (GM) Food

04/21/23 17

What is Genetic Engineering? “Genetic engineering is the technology

for modifying the genetic information in a plant, animal or human in order to produce some desired trait or characteristic”

04/21/23 18

Genetic Engineering VocabularyA. Restriction Enzymes “molecular

scissors” are enzymes that cut DNA only at particular sequences.

B. Plasmids are small circles of DNA found in bacteria.

C. Plasmids are used to replicate a recombinant DNA.

D. Vector - A vector is a small piece of DNA used to carry a gene of interest.

04/21/23 19

Genetic Engineering: Recombinant DNA Technology

04/21/23 20

Restriction Enzymes are Enzymes that Cut DNA Only at Particular Sequences

04/21/23 21

The enzyme EcoRI cutting DNA at its recognition sequence

Different restriction enzymes have different recognition sequences.

DNA Cut by a Restriction EnzymesCan be Joined Together in New Ways

04/21/23 22

These are recombinant DNAs and they often are made of DNAs from different organisms.

Plasmids are Used to Replicate a Recombinant DNA

04/21/23 23

A. Plasmids are small circles of DNA found in bacteria.B. Plasmids replicate independently of the bacterial chromosome.

Human Insulin Production by Bacteria1. Isolate human cells and grow them in tissue culture.

2. Isolate DNA from the human cells.

3. Isolate plasmid DNA from a bacterium.

4. Use the same restriction enzyme to cut the plasmid DNA.

5. Mix the recombinant plasmid with bacteria.

6. Allow the new bacteria to incorporate the recombinant plasmid into the bacterial cell.

7. Grow trillions of new insulin producing bacteria (this is when cloning takes place).

8. A fermentor is used to grow recombinant bacteria.

9. Collect the bacteria, break open the cells and purify the insulin protein.

04/21/23 24

Harnessing the Power of Recombinant DNA Technology – Human Insulin Production by Bacteria

04/21/23 25

04/21/23 26

Human Insulin Production by Bacteria

6) join the plasmid and human fragment

and cut with a restriction enzyme

Human Insulin Production by Bacteria

04/21/23 27

Mix the recombinant plasmid with bacteria.

Screening bacterial cells to learn which contain the human insulin gene is the hard part.

Route to the Production by Bacteria of Human Insulin

28

One cell with the recombinant plasmid

A fermentor used to grow recombinant bacteria.

This is the step when gene cloning takes place.The single recombinant plasmid replicates within a cell.

Route to the Production by Bacteria of Human Insulin

29

The final steps are to collect the bacteria, break open the cells, and purify the insulin protein expressed from the recombinant human insulin gene.

04/21/23 30

Route to the Production by Bacteria of Human Insulin.

Overview of gene cloning.

Genetic Engineering: Insulin Production Overview

04/21/23 31

Monday (March 12, 2012)Genetic Engineering

Journal Question: What is a plasmid?

*1. Lecture II: Genetic Engineering

2. Comprehensive Exam next Monday.

04/21/23 32

Reviewing Genetic Engineering

04/21/23 33

Banking GenesA. The massive Svalbard Global

Seed Vault is built into the permafrost deep in a mountain on a remote arctic island in Norway

04/21/23 34

Banking on GenesB. Built in 2008

After receiving its first deposits, a “doomsday” seed vault on an Arctic island has amassed half a million seed samples, making it the world’s most diverse repository of crop seeds.

04/21/23 35

Most Widely Used Genetically Modified Crops are…

1. Cotton plants with a built-in resistance to insects.

04/21/23 36

Most Widely Used Genetically Modified Crops are…

2. Corn and Soybeans resistant to the herbicide Roundup.

a. Allowing Farmers to employ no-till techniques to farming.

04/21/23 37

Which country is the leader in plant biotechnology?

Answer: China

A. They have recently sequenced the rice genome.

04/21/23 38

Problems with Genetic Engineering Technology1. Environmental Problems

2. Food Safety

3. Access to the New Techniques

04/21/23 39

Environmental Problems

A. Pest –resistant properties of transgenic crops.

B. If pests have a broad exposure to the toxin or some other resistance incorporated into the plant, it is possible that they will develop resistance to the toxin and thus render it ineffective as an independent pesticide.

04/21/23 40

Food Safety

A. Food safety issues arise because transgenic crops contain proteins from different organisms and could trigger an unexpected allergic response to people who consume the food.

04/21/23 41

Access to the New Techniques

A. Relates to the developing world.

B. Farmers in the developing countries are unable to afford the higher cost of the new genetically altered seeds.

04/21/23 42

Other Types of Genetic Engineering: 1. Transgenic Engineering

a. Putting genetic information from one type of plant or animal into another.

2. Cloning

a. Making exact genetic copies of an existing plant or animal.

04/21/23 43

Transgenic Organisms

A. An organism is called “transgenic” if it has genetic information added to it from a different type of organism.

B. Viruses do something of this sort when they infect plants, animals or humans.

C. Humans have begun to do this with plants and animals.

04/21/23 44

Transgenic Organisms D. This is the work that is furthest

along:(1) Corn with its own insecticide.(2) Soybeans & cotton resistant to herbicides.(3) Papayas resistant to viruses

04/21/23 45

Transgenic OrganismsF. Human genes have been

inserted into: (1) Bacteria (Prokaryotes) (2) Mice

G. To produce various human proteins for treating diseases.

04/21/23 46

Making Transgenic Mice

04/21/23 47

Advantages of Transgenic OrganismsA. Plants:

(1) More disease-resistant.(2) Larger yields.(3) More transportable.

B. Animals:(1) Make proteins for medicinal

purposes.(2) Make organs for transplant

to humans.

04/21/23 48

Cloning: “Exact Copies”

A. A “clone” is an exact copy.

B. In genetics, a clone is a genetic copy of another organism.

C. Clones occur naturally: Asexual breeding in plants & lower animals Identical twins (triplets) in higher animals

D. For centuries it has been known that simple animals – worms & starfish – can be cloned by cutting them in half.

E. This doesn’t work for higher animals!

F. Part of the problem is cell specialization: Nerve, Bone, Muscle, etc.

04/21/23 49

Cloning in the 20th CenturyA. We now realize that each specialized

cell has all the genetic information, but much of it is turned off.

B. Problem – how to reset the “program” so this information is usable?C. Cloning of frogs successful in 1950sD. Cloning of livestock from fetal cells

in 1970s.

04/21/23 50

Cloning in the 20th Century

E. The human genome (an organism’s genetic material) consists of 3 billion base pairs of DNA and about 30,000 genes.

(1) 97% of our DNA does not code for protein product.

-mostly consisting of repetitive sequences that never get transcribed.

04/21/23 51

Cloning in the 20th Century: Hello Dolly

F. Clone from an adult sheep cell by Scots researchers under Ian Wilmut.

G. Had only one success in 300 attempts.

H. Dolly grew to maturity, and successfully

had a lamb by natural means in 1998.I. But Dolly seems to be prematurely old.

04/21/23 52

Cloning

04/21/23 53

Cloning

04/21/23 54

04/21/23 55

Genetic Engineering

Genetic engineering (also known as genetic manipulation or GM) is not the same as cloning.

-Though cloning techniques are used in genetic engineering, the two processes should not be confused.

04/21/23 56

Genetic Engineering versus CloningA. Cloning:

1. Produces exact copies2. Genes replicated within the

same species.B. Genetic Engineering:

1. Produces a totally unique set of genes.

2. Genes can be swapped across species.

04/21/23 57

Selective Breeding versus Genetic Engineering

A. In the past, humans have brought about change in the genetic make-up of organisms by means of selective breeding (artificial selection) i.e. Purebreds

B. Genetic engineering brings about such change by scientifically altering an organism's genetic code.

04/21/23 58

Genetic Engineering Overview

1. In genetic engineering enzymes are used to cut up and join together parts of the DNA of one organism, and insert them into the DNA of another organism.

2. In the resulting new organism the inserted genes will code for one or more new characteristics - for example producing a new substance, or performing a new function. The organism has been genetically re-engineered

04/21/23 59

Other names for Genetic Engineering:A. This technique is also known as

gene splicing or recombinant DNA technology (because the DNA is recombined in the vector molecule.

B. Vector - A vector is a small piece of DNA used to carry a gene of interest. Besides the gene being studied, a vector may contain elements which are used to help the gene integrate into a genome.

04/21/23 60

Why does genetic engineering work?A. Genetic engineering works

because there is only one code for life. The set of instructions for which a gene is responsible work whichever organism the gene is in, and whatever instructions that gene gives are carried out within the cells of the recipient.

B. Theoretically the possibilities are limitless, although this sort of manipulation gives rise to strong feelings for and against.

04/21/23 61

Applications of Genetic EngineeringA. One field in which genetic engineering

has had a huge impact is the mass production of insulin to help diabetics. Scientists have isolated the gene responsible for making human proteins, including the insulin hormone. This gene is inserted into the bacterial DNA, and the microbes then clone themselves rapidly, making identical copies of themselves, all with the new gene and all capable of making human insulin.

B. This is a cheap way of producing sufficient quantities of exactly the right hormone, for everyone who needs it.

04/21/23 62

Other Applications of Genetic EngineeringC. Producing interferon, a human

protein which stops viruses multiplying inside the body.

D. Producing human growth hormone to treat growth abnormalities

E. Blood clotting factor to treat hemophiliacs.

F. Used in industry to produce enzymes for use in biological washing powder.

G. Producing pest resistant crop varieties.

H. Producing tomatoes and other produce that stay fresh much longer.

04/21/23 63

Introduction to Genetic Engineering 1. With genetic engineering

scientists directly manipulate genes.

a. It frequently involves the use of recombinant DNA, which is composed of DNA segments from at least two different organisms.

04/21/23 64

Commercial Applications1. An example is the use of

recombinant DNA technology to make interferon, a virus-destroying protein naturally produced by the human body.

04/21/23 65

Production of Synthetic Interferon Involves:1. Isolating the human gene that codes

for the interferon production.2. Splicing this gene into a strand of

bacterial DNA.3. Inserting recombinant DNA into a

bacterium.4. Cloning the bacterium and collecting

the product: Interferon.

04/21/23 66

Isolation of a GeneA. The first step in the process is

isolating the human interferon gene.

B. Genetic engineers use restriction enzymes, proteins that cut a DNA molecule into pieces.

04/21/23 67

Isolation of a Gene

C. The restriction enzyme EcoRI cuts DNA wherever the sequence C-T-T-A-A-G occurs.

D. Other restriction enzymes cut DNA at different nucleotide sequences.

E. By using the proper restriction enzymes scientists can cut the human interferon gene out of its chromosome.

04/21/23 68

Gene Splicing

04/21/23 69

Isolation of a GeneF. Once the gene for

interferon is removed, it is separated from the rest of the DNA and then inserted into a strand of bacterial DNA.

04/21/23 70

Gene Splicing

A. Gene splicing is the process by which a gene from one organism is placed into the DNA of another organism.

B. The human interferon gene is placed into the DNA of E. coli, the common bacterium of the human intestine.

04/21/23 71

Gene Splicing C. In addition to a single, circular

chromosome, E. coli contains a single, small ring of DNA called a plasmid.

(Plasmid = a single ring of DNA in bacteria)

D. Human DNA is inserted into this plasmid.

04/21/23 72

Gene Splicing E. The plasmid ring is removed

from the bacterium and the opened with a restriction enzyme.

F. The human interferon gene and the bacterial plasmid have “sticky ends” –unpaired bases at each end of the DNA segment, where they were cleaved by restriction enzymes.

04/21/23 73

Gene SplicingG. As the human DNA is spliced

into the plasmid DNA, the unpaired bases of each bond readily.

H. Consequently, a newly formed plasmid contains both human and bacterial DNA.

04/21/23 74

Insertion, Cloning, and Collecting A. Once a DNA fragment is

incorporated into a plasmid, the plasmid is inserted into another bacterium, which is then placed in a culture medium, where it divides and replicates.

04/21/23 75

Insertion, Cloning, and Collecting B. Each time a bacterium divides a

new copy of the plasmid DNA, which includes the human DNA gene, is created.

C. This process by which the human gene is replicated is called gene cloning

04/21/23 76

Insertion, Cloning, and CollectingD. Because E. coli can divide every

20 minutes, gene cloning is an efficient way to produce many copies of a specific genetic sequence.

04/21/23 77

Insertion, Cloning, and CollectingE. The gene for human interferon is

thus expressed in bacterial cultures and the resulting interferon protein is collected and eventually used by physicians.

04/21/23 78

04/21/23 79

Genetically Modified (GM) Crops

The ear of genetically engineered corn at top contains a toxin that kills worms.

04/21/23 80