Genetic Engineering.pptx - MHSBioMrP2010

Genetic Engineering

Chapter 10

What we are hoping to accomplish

•
Define genetic engineering.

•
Explain the principles of genetic engineering.

•
Explain and use the tools of genetic engineering.

•
Show and describe the practical uses of genetic
engineering.

•
Discuss the various steps in PCR and
transformation.

Biotechnology
-
Genetic Engineering

•
The application of biological and physical
principles to alter the genes of organisms to help
solve human problems

▫
Improving crops and their yield

▫
Producing insulin in bacteria

▫
Genetically modified vaccines

▫
Improving the freshness of fruits and vegetables


Biotechnology

Historical
Aspects

Plant breeding

Animal Breeding

Wine making

Dairy products

Bread

Antibiotics

Herbal medicine

Business and
Industry

Biomedical

Agriculture

Forensics

Industrial
production

Conservation

Wildlife
management

Multidisciplinary
Research

Biochemistry

Molecular genetics

Biophysics

Immunology

Cell culture

Fermentation

Bacteriology

Bioengineering

Genetic medicine

Conservation

Evolutionary
biology

Social and Ethical
Implications

Genetic privacy

Ownership of info.

Screening

Engineered food

Insurance

Treatment

DNA profiling

Patenting of life

What does genetic engineering mean?

•
It means to isolate a gene from the DNA of one
organism and transfer the gene into the DNA of
another.


•
We can do this because DNA is DNA in all
organisms. It is made out of the same 4 nitrogen
bases. The only difference is the sequence.


•
From there, DNA codes for RNA and proteins.

Recombinant DNA

•
A molecule of DNA made from pieces of DNA
from separate organisms.

Restriction Enzyme

•
An enzyme that is used to “cut” DNA at certain
places along
BOTH

sides of the DNA.


•
Most commonly used Restriction Enzymes

▫
BamHI
-

G/GATTC

▫
HindIII
-

A/AGCTT

▫
HpaII
-

C/CGG

▫
EcoR1
-

G/AATTC

Plasmids

•
Circular piece of DNA found in bacteria.

▫
Replicates separate chromosome DNA

▫
Contains very few genes

▫
Useful because they have antibiotic resistance
genes on them.

▫
Typically used as
VECTORS

to transfer DNA
from one organism to another.

Polymerase Chain Reaction (PCR)

•
A way to make an unlimited number of copies of
one gene.


•
How? DNA Polymerase is used to replicate the
same DNA segment over and over.


•
Why?

▫
Increasing the number (Crime Scenes)

▫
Sequencing (Prenatal diagnosis)

▫
Cloning (Woolly Mammoth)

3 Steps to PCR

1.
Denaturing
-

Stands of DNA separate by
heating

2.
Annealing
-

Reaction cooled, primers bind

3.
Elongation
-

Reaction heated again, DNA
polymerase replicates the rest of the DNA from
primer.


•
Repeat steps 1
-
3 about 20
-
30x

DNA Fingerprinting

•
DNA is sliced into fragments called RFLPs.


•
DNA is then separated by sequence length by gel
electrophoresis.


•
The result is for an individual different lengths
of DNA that were cut by restriction enzymes that
varies among alleles and individuals.

Gel Electrophoresis

How the Gel Works?

•
DNA is negative


•
When DNA is placed next to a negative charge it
will want to move towards the positive charge.


•
The heavier strands will not travel as far as the
lighter strands.

DNA fingerprinting in a crime scene

Who did it? How Do We Know?

Crime Scene

Suspect 1

Suspect 2

Suspect 3

Suspect 4

Use in Crime Scenes

•
3 different Fingerprints are run


•
1/10,000 chance of matching an individual


•
Two fingerprints will only match 1 out of 10,000
X 10,000 times


•
Three matching prints would then be 1 in one
trillion


The DNA

•
Magnified using PCR


•
From blood, bone, flesh, hair follicle, semen,
saliva, any source of DNA


•
Analysis does not compare genes rather the
information between genes where a great
amount of variation exists

▫
RFLP analysis


Restriction Fragment Length
Polymorphism (RFLP) Analysis

•
Identifies locations and numbers of restriction
sites of DNA.

Human Genome Project

•
Purpose: Sequence the entire human genome.

•
Why: detect, treat, and prevent genetic diseases.

•
Completed in 2003

•
Some of their findings for human DNA

▫
Approximately 20,000
-
25,000 genes
(23,000)

▫
3 billion base pairs make up the DNA.

▫
Only 1.5% is actually coding for protein

•
Still left to do? Figure out how it all fits together.

Typical Transformation Experiment

1.
A target gene is isolated and removed by a
restriction enzyme
.

2.
Another restriction enzyme will cut the DNA of
a
vector

(
plasmids
) and insert our target
gene.

3.
Transfer the vector to the organism we want to
modify.

4.
Transformation
-

the gene becomes one with
its host and replicates producing clone copies.

5.
Screen the cells and select modified organism
by identifying our gene of interest.

Why do we care?

Genetically Engineered Drugs

Genetically Engineered Vaccines

•
Humulin

-

Diabetes

•
HGH
-

Dwarfism

•
Factor VIII, Factor IX
-

Hemophilia

•
t
-
PA
–

Heart attack, clot
buster

•
Lactoferrin

–

Milk protein


•
Herpes II

•
Hepatitis B

•
Influenza


Why do we care?

Cures for Genetic Diseases

Forensic Science

•
Cystic fibrosis

•
Genetic testing

•
Therapy

•
Karyotyping


•
DNA fingerprints

▫
Crime

▫
Paternity

▫
Predisposed to disease


This may involve using stem
cells, by taking somatic cells
(like skin cells) and
dedifferentiating them down
to
pluripotent

stem cells.

Why do we care?

Plants

Animals

•
Ti plasmid causes plant
tumors

•
Glyphosate

resistant crops
(round
-
up ready)

•
Pest resistance

•
Improve taste and nutrition

•
Vaccines grown in plants


•
BST

•
Lactoferrin

•
Cloning