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Skin Test

Ancient Chinese is performing


variolation.

Edward Jenner is vaccinating


cowpox.

variolation

Edward Jenner memorial


hall previous house

Edward Jenner(1749
1823)

Louis Pasteur,18221893

Emil Adolf von


Behring 1854-1917

Microbiologist,
France

Bacteriologist, Germany

Paul Ehrlich (1854~1915)

Elie Metchnikoff
(1845~1916)

Lymphocytes(T,
B)

NK cell

eosinoph
il

Dendritic cell

Monocyte/macrop
hage

basophil

Mast cell

Immunocytes

neutrophil

erythrocyt platel
e
et

Immune Response
The whole process including antigen
recognition and antigen elimination .
1 Innate Immune Response non-specific, inborn
barrier
phagocytosis
normal humor component
2 Adaptive Immune Response specific, acquired
humor immunity
cell immunity
mucosa local immunity

Comparison between innate


immunity and adaptive immunity
innate immunity

adaptive immunity

Generated little by little during


long term evolution

Stimulated by antigen

Commonly owned by different


germlines, herediable

Specially owned by some


individuals, non-herediable but
form immune memory

Non-specific (respond
generally

Specific (respond to special


antigen)

React rapidly and generally

Undergo process including


recognition of antigen,expansion
and differentiation and effect

Engulfment
IgG-opsonized
particle

FcR

pseudopods

target
Phagocyte

FcR activation induces


actin polymerization

30s

target
Phagocyte
Extension of pseudopods
around the target

3 minutes

Engulfment
Complement Receptor
Complement-opsonized
particle

Phagocytosis by complement receptor


requires activation with inflammatory
cytokines. FcR does not require activat
target by other signals.
No activation Activation
phagocyte

Phase contrast
target
No pseudopods

Sinking of target
into phagocyte
Staining for CR
CR surrounds the
phagocytosed
particle.

Maturation of Phagosomes
phagocytosis
receptor

Endocytosis
receptor

Clathrin-coated pit

Actin polymerization

phagosome

endosomes

Fusion with endosomes and lysosomes


Gogi

Hydrolytic enzymes
lysosomes
Phagolysosome

Existing TB Vaccine Ineffective

BCG provides unreliable protection against


pulmonary TB, which accounts for most TB
disease worldwide

BCG is not know to protect against latent TB

BCG is not recommended for use in infants


infected with HIV due to increased risk for
severe BCG-related complications

Despite wide use, particularly in high burden


countries, BCG has had no apparent impact
on the growing global TB epidemic

BCG does reduce risk of severe pediatric


TB disease, so it should continue to be used
until a better TB vaccine is available

BCG introduced in
1921

Previous Lesson

DNA and the Human


Genome Project

Question: How many types of


proteins are in a human and
why do we need so many
different types of proteins?

Each humans has about two


million different types of
proteins.
Proteins play a role in hair and
muscles. They are essential to
the immune system and most
hormones and enzymes are
proteins.

Question: Why would a plant


cell have DNA in its nucleus
and ribosomes in its
cytoplasm?

Based on Raven et al., Biology, McGraw HIll

Like animals, plants must produce proteins


to regulate their biochemistry

Question: You are a unique


person. What is the blueprint
for life that makes you
unique, and how does it act
as a blueprint?

DNA is the Blueprint for Life


Each person has a unique complement of DNA
which causes
Each person to have a unique complement of
protein
which contributes to
Each person to have a unique appearance and
behavior

Question: Why did many


scientists think that
sequencing the human
genome was impossible?
What made it possible?

The human genome consists of


about 3 billion letters
(nucleotide bases)
15 billion dollars of federal funding
made it possible

Todays Lesson

Individuality and
Cloning

Overview of Lesson

Basics of genetics
Humans as unique individuals
Identical twins
Cloning of Dolly
Human cloning

Two Brothers

Matt:
Biology
Fishing
Hunting

Bryan:
Religion
Philosophy
Computers

Each human body


cell has 23 pairs of
homologous
chromosomes
(46 total)

Karyotype

Half of the
chromosomes are
from the mother and
half are from the
father

Each member of a homologous pair has the same genes but


the alleles (specific expression of each gene) on each member
might be the same or different

Widows
peak
allele

Gene
for
hairline

Straight
hair
allele

Alleles are represented by letters

Widows Peak (W)


is a dominant allele

Straight Hairline
(w) is a recessive
allele

http://images.google.co

Meiosis: formation of gametes in gonads

Based on: Mader, S., Inquiry Into Life, McGraw-Hill

Meiosis results in:


gametes with half the number of
chromosomes as the number in our
body cells - 23 (n) instead of 46 (2n)
gametes with different chromosome
combinations

Overview of Lesson

Basics of genetics
Humans as unique individuals
Identical twins
Cloning of Dolly
Human cloning

Why are humans such unique


individuals?
because each of us has a unique set of
genetic instructions (set of alleles in DNA
of chromosomes) due to the random
assortment of chromosomes during
generation of gametes (eggs and sperm)
through meiosis

During meiosis in humans


chromosomes can line up 223 or
10 million (107) different ways in
meiosis I
so each person can produce 10
million (or 107) different types of
gametes

For each couple


107 types of sperm
x
107 types of eggs
1014 types of children

Crossing-over between chromatids during


meiosis makes it even more complex

Based on: Mader, S., Inquiry Into Life, McGraw-Hill

The two brothers


represent 2 out of
more than 1014
possible children that
a couple could
conceive
They are genetically
programmed to be
different

Overview of Lesson

Basics of genetics
Humans as unique individuals
Identical twins
Cloning of Dolly
Human cloning

Identical twins are produced when the two


cell stage splits, forming two separate cells.
Then the cells continue to divide separately
forming two embryos.
Two cell stage

Based on: Mader, S., Inquiry Into Life, McGraw-Hill

Identical Twins
Identical chromosomes (DNA)
Identical proteins
Very similar appearances

Identical Twins

http://images.google.com/

Conjoined twins joined at


the head

Inner cell masses subdivide


but do not separate
completely, forming conjoined
twins

http://www.youtube.com/watch?v=BkKWApOA

Overview of Lesson

Basics of genetics
Humans as unique individuals
Identical twins
Cloning of Dolly
Human cloning

What is cloning?
Taking an adult (fully differentiated)
cell, and restarting its genetic program
and protein synthesis
(as if it were a fertilized egg)

Why was cloning thought to be


impossible?
as the genes of DNA regulate protein
synthesis during development, the
genes are turned off
(genes active at the zygote stage
are inactive in the adult)
to clone a mammal, the DNA would
have to be turned back on, as if being
read from the start (zygote stage)

In slide show mode,


use the hot link
below to download
a copy of the cover
of Time Magazine
on Dolly

TIME Magazine Cover: Dolly, Cloned Shee


p - Mar. 10, 1997

Born in Scotland,
Dolly was the
first mammal to be
cloned (1997)
This showed that
DNA in adult cells
can be re-started

Ethical
Issue
of the Day #2

Stem Cells

What are stem cells?


What are they good for?
What is an embryo?
Do all stem cells used in research come from
embryos?
Why are embryos useful to make stem cells?
Will banning stem cell research put an end to
the creation and destruction of human
embryos in test tubes?

2004 Lee Bardwell

What about cloning


animals?
Genetic Savings & Clone, Inc.
www.savingsandclone.com

clone

donor

Cloning of Dolly

Based on: Raven, et al., Biology, McGraw-Hill

Cloning of Livestock
Sheep
Goats
Mice
Cows
Pigs
U.S. News and World Report, March 27,
2000
U.S. News and World Report, December 21,1998

High Failure Rate in Animals


2% success rate
Miscarriage frequent
Animals die after birth
Some animals have developmental
abnormalities
Death of offspring is 10 times
higher than normal before birth
and 3 times higher after birth

Overview of Lesson

Basics of genetics
Humans as unique individuals
Identical twins
Cloning of Dolly
Human cloning

Why would the U.S. House of


Representatives vote in 2001 to
ban human cloning?

Because of the unacceptable


high risks of miscarriages and
birth defects associated with
cloning
Furthermore, most Americans are
opposed to human cloning

What would be the difference


in a cloned child and its
parental clone?
What problems might this
create for the cloned child?

A cloned child would be the


genetically identical twin of the
parental clone, but have a different
personality (because personality
emerges from nature and nurture)
Parents might place unrealistic
expectations
An exception might be cloning a child
who died at birth

What is the difference


between therapeutic cloning
and reproductive cloning of
humans?

Therapeutic cloning could be used to


produce embryos with stem cells that could
be used to treat diseases such as
Parkinsons Disease and diabetes. The cells
would be a perfect genetic match for the
person donating the cells.
Reproductive cloning will be used to
produce embryos that will be implanted into
a womans uterus and allowed to develop
into an infant.

Next Lesson

Genetic Disorders and


Gene Therapy

Signature Program:
Reproductive Sciences

Reproductive Sciences Laboratory


(RSL)
Faculty
Mark Westhusin
Duane Kraemer
Charles Long
Visiting Scientist
Mundhir Ridha
Staff
Taeyoung Shin
Christy Bormann

Graduate Students
Carrie Hanna
Jie Liu
Arantza Lassala
Jessica EppleFarmer
Abigail Casillo
Lara Charles
Beth Kidd-White
Suzanne Mengas
James Williams

Affiliations
Interdisciplinary Faculty of Reproductive
Biology (IFRB)
42 Members, 4 Colleges
11 CVM, 6 Joint Appointments in CVM
Center for Genomics and Biotechnology

Nucleus Transfer
Collecting and storing nucleus
donor cells

Surgical procedure
Live animal
Post mortem

Culturing cells
Cryopreservation (freezing) of cells

The Technology
Preparing nuclei for transfer

Regulating cell cycle


Serum starvation

Obtaining recipient ova

In vivo matured
In vitro matured

The Technology

Micromanipulation

Nucleus Transfer

Enucleating ova
(removing the
nucleus)

The Technology

The Technology

Transfer of the cell

The Technology

The Technology

Cell fusion

Cell Fusion

Cell Fusion

The Technology

Alternative Method
Intracytoplasmic
nucleus injection

The Technology
DNA Reprogramming

Methylation
Demethylation

The Technology

Activation

Electrical
Chemical
Insitol 1,4, 5-triphosphate
Ionomycin + b-dimethylaminopurine

Examples of Clones
Second Chance
21 year-old Brahman Bull

Photos courtesy of: Hill, et al., Biology of Reproduction 62:1135 (2000)

Second Chance (3 years old)

Examples of Clones
86 Squared

Brucellosis
resistant bull
Tissue frozen
for 15 years

86 Squared (2 years old)

Examples of Clones

White-tailed Deer (Dewey)

Dewey 16 months old

Examples of Clones
CC The first
cloned cat
Progenit
or

Current Research Projects:


In Utero stem cell trasplantation to correct
hemophilia A. NIH subcontract through
University of Nevada, Reno, $680,500. P.I.
of Subcontract: D.C. Kraemer, 2003-2007

Objective:
To re-establish and extinct strain of
hemophilic sheep from 6 straws of frozen
semen
Work done at U.N.-Reno (mobile lab)

Full Spectrum of Reproducive


Technologies
Multiple Ovulation Embryo Transfer
(MOET)
Artificial Insemination (AI)
In-Vitro Oocyte maturation (IVM)
In-Vitro Fertilization (IVF)
Intra Cytoplasmic Sperm Injection (I CSI)
Nucleus Transfer Cloning (NTC)

Rehearsal Experiment
Procedures
MOET
MOET+NTC
IVF
ICSI
NTC

Transfers
4
4
3
2
2
15

Pregnancies
3
2
0
0
0
5

First Experiment
Procedures
MOET
IVF
ICSI
NTC

Transfers
0
0
17
2
19

Pregnancies
0
0
11
0
11

Research Projects
Somatic cell nucleus transfer in the
canine. Texas Heart Institute. $72,721. P.I.
D.C. Kraemer, 2004-2005

Canine Cloning
9 bitches

Huntsville (Labradors and hounds)


Local (Basset x Dalmatian)

Procedure
Oocyte collection and in vitro
maturation (IVM)

Results to Date
IVM- collected average 64 MI and MII
Transfer average 36 couplets
No full term pregnancies so far

http://highered.mcgrawhill.com/olc/dl/120078/micro10.swf

II. Historical Background

A. 1953 Watson and Crick: DNA Structure Predicts a Mechanism of Replication


It has no escaped our notice that the specific pair we have postulated immediately suggests
a possible copying mechanism for the genetic material.

B. 1958 Meselson and Stahl: DNA Replication is Conservative

The Meselson-Stahl Experiment


the most beautiful experiment in biology.

All
hybrids
1/2 old:
1/2 new

1/4 old:
3/4 new

1/2 hybids:
1/2 new
All
hybrids

Three potential DNA replication models and their predicted


outcomes

The actual
data!

III. General Features of DNA Replication

DNA Synthesis:
1. requires a DNA template and a primer with a 3 OH end.
(DNA synthesis cannot initiate de novo)
2. requires dNTPs.
3. occurs in a 5 to 3 direction.

Short RNA molecules


act as primersin vivo

I. SRY codes for Testis-Determining Factor

Was the object of an intense search.

SRY gene on the Y chromosome was identified as the gene that codes
for TDF:
SRY is translocated to X in rare XX males
SRY is absent from Y in rare XY females

The home run experiment by Koopman et al. used transgenic mice.

III. Ch. 10 - chromosome mutations


Prelude to nondisjunction
Terminology to describe variations in
chromosome number

Aneuploidy: 2n + or - chromosomes
Monosomy: 2n - 1
Trisomy: 2n + 1

Euploidy: multiples of n
Diploidy: 2n
Triploidy: 3n
Tetraploidy: 4n

Nondisjunction of X chromosome

Nondisjunction also occurs with autosomes

Partial Monosomy:
Cri-du-chat Syndrome (46, -5p)

Mental retardation, abnormal development of glottis and larynx


1 / 50,000 live births

Trisomy:
Down Syndrome (47, +21)

Characteristic facial patterning, mental retardation


1 / 800 live births

Down Syndrome Characteristics

Most often occurs by nondisjunction of chr. 21 during meiosis; in theory could


occur in either mom or dad, but 95% of these trisomies have defective egg as
source

Prone to respiratory diseases, etc.

About 30% of all cases of mental retardation in U.S.

1/25 can read; 1/50 can write

Detectable by amniocentesis or chorionic villus sampling (CVS)

Incidence of Down Syndrome Increases


with Maternal Age
All eggs are formed by
birth and arrested in
meiosis; is the
correlation of increased
age and the syndrome
due to more
nondisjunction in older
eggs?

Other trisomies:
Patau Syndrome (47, +13)

1 / 19,000 live births

Edwards Syndrome (47, +18)

1 / 8000 live births

H&J Fig. 4.19

What is Human Cloning?


Human embryo clone means a human
embryo that is a genetic copy of another
living or dead human, but does not include
a human embryo created by the
fertilisation of a human egg by human
sperm.
(s.8 Prohibition of Human Cloning Act 2002)

What is Human Cloning?

Method 1: Usual reproduction [in utero or in vitro]


Method 2: Somatic Cell Nuclear Transfer [SCNT]

What is Human Cloning?

Cellular Component
where a gene product acts

24th Feb 2006


Lomax

Jane

Cellular Component

24th Feb 2006


Lomax

Jane

Cellular Component

24th Feb 2006


Lomax

Jane

,
;
,


.
,


.
,

Polymerase Chain Reaction (PCR)

Machine that allows us to


perform PCR.
PCR

http://academic.brooklyn.cuny.edu/biology/bio4fv/page/genetic-engin/pcr2484.JPG


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