Biotechnology

Vince

Zi

Shan

Zach

Verdi

Fixler

Most of the organisms

that are used in
agriculture

(crops,
livestock) have been
produced by human

beings by selective
breeding, which is
sometimes called
artificial

selection.

Humans have been
doing this for
thousands of years!
For example, the
wild
banana

looks
very different from
the yellow
Chiquita

you might purchase
in the store.

These fruit are
not

sweet

and are filled
with unpleasant
seeds
.

The good
-
tasting
varieties

most of us
take for granted were
produced long ago by
Africans

who crossed
different
varieties
, and
selected the offspring
that were more
pleasant to taste in
every generation.

The results of
these crosses,
called
hybrids,

became the basis
of the modern
fruit.

In the same way,
Native
Americans

selected varieties of a
naturally
-
occurring
plant called
teosinte

that were a little bit
taller and had
more

seeds
.

Over time, this plant,

known as
maize

or
corn, became very tall
and produced a
gigantic cob with many
kernels
.

The same thing is true for most
varieties of livestock and
pets
:
their
populations

look different
from those in nature.
Why?
Because, over time,
humans

have
selected for the
trait

that they
find most useful or attractive. To
do this, they cross individuals with
similar characteristics, a process
known as
inbreeding
.

Woof!

What about

animals?

The same thing is true for most
varieties of livestock and
pets
:
their
populations

look different
from those in nature.
WHY ?

Because, over time,
humans

have
selected for the
trait

that they
find most useful or attractive. To
do this, they cross individuals with
similar characteristics, a process
known as
inbreeding
.

This is not

necessarily in
the animal’s
best interest,
because it can
increase the
chances of
offspring having
genetic
disorders.

For example,
Dalmation

females often
have pups born
deaf

in one ear,
or with

liver

disease.

Genetic

engineering is
really no different! It’s
just another form of
selective breeding, only
in this case humans are
selecting the
genes
directly, instead of the
traits

based on the
genes
.

Grafting


•
Go to computers and see two ways to
clone… Should take no more than 10
minutes!

•
You will be quizzed on what you learn in
the next 10 minutes!

Once scientists learned
that DNA was the
genetic
material, they
developed techniques to
harvest

it from living
cells.

Restriction Enzymes

(R.E.)

•
*used to cut virus DNA, into fragments to
kill the virus(defense mechanism).

•
R.E. are found in bacteria

•
*large variety of restriction enzymes
exist:


http://www.youtube.com/watch?v=8rXizmLjegI


•
Bam HI G/GATCC

•
Hind III A/AGCTT

•
EcoRI

G/AATTC

Scientists do more than
just collect DNA,
however. They
concentrate and
purify
it, then cut the long
strands of DNA into
smaller fragments with
restriction

enzymes
made by bacteria.

There are hundreds of

different
restriction

enzymes known, and

each one cuts the DNA

at a specific
nucleotide
sequence .




.

Restriction enzyme
EcoRI cuts the DNA into
fragments.

DNA sequence

recognition sequence

“sticky end”



For example, the enzyme
EcoRI

cuts the sequence


‘G/AATTC’ between the ‘G’ and the first ‘A’.

This leads to many DNA fragments of different
sizes

.



.



.

•

uses a
power

source,


a tray filled with
buffer


solution and an
agarose


gel


•

involves ‘loading’ a


mixture of DNA fragments


into little cavities in the


gel, called
wells



.



.

•

applies an electric field


to the
negatively
-
charged


DNA fragments, which


move to
positive pole.


•

separates the fragments


over time, since the


smaller

fragments move


faster and
farther



.



.

•

produces a pattern of


lines that can be read


like a
bar
code at the


supermarket:

Click me to see

a Flash animation!



.



.



.



.

DNA plus
restriction
enzyme

mixture
of DNA
fragments

agarose
gel

power
source

longer
fragments

shorter
fragments

Heating destroys
most enzymes,
including the enzyme
that helps DNA

copy itself, DNA

polymerase
.


But in 1969, a

microbiologist named

Thomas Brock discovered

a species of bacteria

living in the hot springs

of Yellowstone National

Park . . .



This bacteria, which

Brock named
Thermus

aquaticus
, thrives at

just below the

boiling

point of water. . .


Researchers use
this enzyme to
make many copies
of the DNA with a
technique called
polymerase chain
reaction
, or PCR.

PCR takes advantage

of the fact that the
double
-
stranded DNA
fragments will
separate

into two
single
-
stranded
fragments when heated.

In PCR, researchers will

heat the DNA along with

the
Taq
enzyme, along

with a vast surplus of all

the different
nucleotides
:

G’s, C’s, A’s and T’s.

The strands
will separate!


(completely)

The mixture is then

allowed to cool, and

as it does the enzyme

will attach the free
nucleotides

to the
exposed strands . .

. . leading to
two

copies

of double
-
stranded DNA!

This process of

heating and cooling

can be repeated

several times, each

time doubling the

number of copies: in

less than 12 hours,

researchers can

manufacture
millions

of copies of a single

strand.

DNA fragment
to be copied

DNA heated to
separate strands

DNA polymerase adds
complentary strands

DNA fragment
to be copied

DNA heated to
separate strands

DNA polymerase adds
complentary strands







PCR

cycle: ____ _____ _____ _____ _____



DNA

copies: ____ _____ _____ _____ _____

1


2

2


4

3


8


4


16


5


32

laser
Once strands are copied,

researchers will label

the DNA by attaching
different
-
colored chemical
dyes to the different
nucleotides
.

O
nce

s
tr
and
s

a
re

cop
i
e
d,

re
sea
r
ch
ers

wi
ll

la
be
l

th
e

D
NA

by

att
ach
in
g

di
ffe
r
ent
-
col
or
ed
ch
e
m
ic
a
l
dy
es

t
o

th
e

d
if
fer
en
t

nu
cle
o
tid
es
.


Si
nce

e
ac
h ba
se

i
s

labe
l
e
d

Wi
th
a

d
iffe
r
ent

co
lo
r,

th
e

o
rder

of

the

G’s
, T’
s

an
d C’
s
ca
n be

r
ead
b
y
a

laser

and

r
ec
ord
ed

b
y
a

co
mpu
ter
.


laser

The success of the Human
Genome

Project is based

on the fact that the entire process is
automated


Gel
electrophoresis

and
DNA
sequencing

can be
used to identify individuals
(
DNA fingerprinting
) or to
develop genetic profiles of
entire
populations
.




A famous photograph from the O.J. Simpson trial: the blood allegedly


found on the famous athlete’s sock was not his!



Gel
electrophoresis

and
DNA
sequencing

can be
used to identify individuals
(
DNA fingerprinting
) or to
develop genetic profiles of
entire
populations
.

Click on the map!

This information can
be used to determine
paternity
, to link
suspects in a
criminal
investigation to a
crime

scene or to
probe an individual’s
medical

history or
ancestry.


In many cases, it is not necessary to have a copy

of all of an individual’s DNA, only a specific

sequence

which is already well
-
described.


In many cases, it is not necessary to have a copy

of all of an individual’s DNA, only a specific

sequence

which is already well
-
described.


Not all parts of the DNA

are
expressed (exons),

and

many parts that are
not

expressed (so
-
called
‘junk’
DNA
, or
introns
) have
sequences that are repeated
over and over.

These regions of DNA accumulate many harmless

mutations
which are different for every individual.


So, when these regions are cut by
restriction

enzymes, they create unique patterns that can be

used to identify an individual.



This technique,
often used in law
enforcement,

is called
RFLP
:


R
estriction


F
ragment


L
ength


P
olymorphism


(pronounced ‘rif
-
lip’)

This technique,
often used in law
enforcement,

is called
RFLP
:


R
estriction


F
ragment


L
ength


P
olymorphism


(pronounced ‘rif
-
lip’)