Download The human genome of is found where in the human body?

The human genome of is found where
in the human body?
A.
B.
C.
D.
Nucleus
Ribosome
Smooth ER
Cell membrane
The cellular structure where proteins
are made is called the
A.
B.
C.
D.
Nucleus
Smooth ER
Ribosome
Cell membrane
DNA and Biotechnology
Announcements
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Ch 21- today
Ch 21, 17- Wednesday
Bone & muscle labs- Due Wednesday
Ch 21, 17 online quizzes- Due Friday
Quiz 10- Friday (chs 21, 17)
Lab today- pGLO (DNA transformation)
Lecture Outline
• DNA- Structure, function, and importance
• How DNA works
– The central dogma
– Transcription and Translation
– The DNA code
– DNA replication
The importance of DNA
The DNA double helix is the code of
life
• The blueprint for all
structures in your
body which are
made of protein
• DNA is comprised of
nucleotides
Nulceotides are the monomers of
nucleic acid polymers
• Consist of a sugar, a
phosphate, and a
nitrogen-containing
base
• Sugar can be
deoxygenated
• Bases contain the
genetic information
There are 4 kinds of DNA bases
Adenine always
matches with
Thymine,
Cytosine always
matches with
GuanineHydrogen bonds
hold bases
together
Living things are extremely complex
• Cellular machinery is
sophisticated and required for
life
• Cellular machinery is made
largely of proteins
• Blueprints for all cellular
machinery are contained in
genes
• Genes are inherited from
parents
• Humans have ~30,000 genes
Proteins give living things the variety
of their structures
Protein variety is generated by 1o
structure- the sequence of amino acids
which make the protein
Amino Acids
• Proteins consist of subunits called amino acids
Figure 2.12
How DNA works
• Replication
• Transcription
• Translation
The sequence of DNA bases is the
code for the primary structure of
proteins
All cells require a copy of the genome
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Genome- all the genes of the cell
Human genome is made of DNA
DNA is similar in all cells
Gene- 1 DNA Molecule (+
proteins the genetic information
to produce a single product
(protein)
• DNA replication copies all cellular
DNA
Replication of DNA
Figure 21.2
In vivo, enzymes such as DNA polymerase
make DNA replication happen
The DNA code
Computers use binary digital code
01000011 01101000
01100001 = A
01100101 01100101
01100010 =B
01110011 01100101
01000011 =c
01100010 01110101
01110010 01100111
00100111 = apostrophe
01100101 01110010
Etc.
00100000 01000100
01100101 01101100
• http://www.geek01110101 01111000
notes.com/tools/17/tex
01100101 =
t-to-binary-translator/
cheeseburger deluxe
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How does the DNA code work?
• atggcttcctccgaagacgttatcaaagagttcatgcgtttcaaa
gttcgtatggaaggttccgttaacggtcacgagttcgaaatcga
aggtgaaggtgaaggtcgtccgtacgaaggtacccagaccgct
aaactgaaagttaccaaaggtggtccgctgccgttcgcttggga
catcctgtccccgcagttccagtacggttccaaagcttacgttaa
acacccggctgacatcccggactacctgaaactgtccttcccgg
aaggtttcaaatgggaacgtgttatgaacttcgaagacggtggt
gttgttaccgttacccaggactcctccctgcaagacggtgagttc
=GFP
The DNA
code is
(nearly)
universal
It uses groups of 3
bases (codon)
3 bases = 1 codon = 1
amino acid
And what
are these
U’s for?
RNA is ribonucleic acid
• Ribose sugar is not
deoxygenated
• RNA is singlestranded
• RNA has Uracil, not
Thymine
• There are many
kinds: mRNA, rRNA,
tRNA, siRNA, etc.
RNA can fold back on itself
• Single strand offers
greater flexibility
Kinds of RNA
mRNA
tRNA
The Central Dogma of Molecular
Biology
• DNA RNA Protein
• DNARNA :
Transcription
• RNA Protein:
Translation
When one DNA molecule is copied to make two
DNA molecules, the new DNA contains
1.
2.
3.
4.
5.
A) 25% of the parent DNA.
B) 50% of the parent DNA.
C) 75% of the parent DNA.
D) 100% of the parent DNA.
E) none of the parent DNA.
DNA RNA Protein Trait
The
Universality
of the DNA
code makes
this
possible
Firefly gene (Luciferase) in a tobacco plant
Transcription and Translation
Transcription: DNA RNA
DNA Codes for RNA,
Which Codes for Protein
Figure 21.3
DNA information is transcribed into
mRNA
Note in DNA:
sense strand
vs. antisense
strand
Translation: RNA Protein
tRNA’s carry an amino acid at one end, and
have an anticodon at the other
Amino acid
attachment site:
Binds to a specific
amino acid.
Amino acid
(phenylalanine)
Anticodon:
Binds to codon on mRNA,
following complementary
base-pairing rules.
Anticodon
mRNA
Figure 21.6
The ribosome matches tRNA’s to the
mRNA, thereby linking amino acids in
sequence
tRNA’s add amino acids one by one according to
mRNA instructions until the protein is complete
We would expect that a 15-nucleotide mRNA sequence will
direct the production of a polypeptide that consists of
1.
2.
3.
4.
5.
A) 2 amino acids.
B) 3 amino acids.
C) 5 amino acids.
D) 7 amino acids.
E) 15 amino acids.
Viruses exploit the universality of the DNA code
to take control of cells
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Basic life cycle of Viruses
Viruses are obligate
intracellular parasites
They inject their genetic
material into their host
Host machinery is
commandeered to massproduce virus
Viruses burst host cell to
infect other cells
Virus Structure
• Many viruses contain
only:
– Protein capsid
– Genome (DNA or RNA)
• Some viruses have a
phosopholipid bilayer
envelope
Bacteria are infected by viruses, too
Bacteriophages attacking
bacterial cell
The lytic bacterial life cycle
Bacteriophages
• Infect bacteria
• Cause formation of
plaques on a lawn of
agar in bacteria
Bacteria use restriction enzymes to
defend against viral DNA
Restriction enzymes cut very specific
sequences of DNA
• Hundreds of different
restriction enzymes
have been found
• Named after bacteria in
which they are found
Viruses such as T4 fight back with DNA
ligase
• Ligase glues DNA back
together
• Viral countermeasures
against bacterial cell
defenses
Scientists commandeer these enzymes to
perform DNA manipulations
• Ligase and restriction
enzymes allow any
sequence of DNA to be cut
and pasted at will
• Plasmids, small loops of
bacterial DNA, can be
modified with any DNA
• Because the genetic code
is universal, DNA will be
read in the same way
Bacterium
Gene inserted into
plasmid
Plasmid
DNA
manipulation is at
the heart
of biotechnology
Bacterial
chromosome
Cell containing gene
of interest
Plasmid
Gene of
interest
Recombinant
DNA (plasmid)
DNA of
chromosome
Plasmid put into
bacterial cell
Recombinant
bacterium
Host cell grown in culture
to form a clone of cells
containing the “cloned”
gene of interest
Protein expressed
by gene of interest
Gene of
interest
Copies of gene
Basic
research
on gene
Gene for pest
resistance inserted
into plants
Protein harvested
Basic research and
various applications
Gene used to alter
bacteria for cleaning
up toxic waste
Protein dissolves
blood clots in heart
attack therapy
Basic
research
on protein
Human growth hormone treats stunted
growth
The pGLO plasmid has
• ori- origin of replication
1. GFP- green fluorescent
protein
2. bla- beta-lactamase
(confers ampicillin
resistance)
3. araC- Arabinose regulator
protein (regulates GFP
expression)
1. GFP: Green Fluorescent Protein
•
discovered in 1960s by Dr. Frank
Johnson and colleagues
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closely related to jellyfish aequorin
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absorption max = 470nm
•
emission max = 508nm
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238 amino acids, 27kDa
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“beta can” conformation: 11
antiparallel beta sheets, 4 alpha
helices, and a centered chromophore
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amino acid substitutions result in
several variants, including YFP, BFP,
and CFP
40 Å
30 Å
2008 Nobel Prize- GFP
• GFP mice
GFP can be fused to cellular proteins
Using GFP as a biological tracer
http://www.conncoll.edu/ccacad/zimmer/GFP-ww/prasher.html
With permission from Marc Zimmer
GFP fusions are useful in Biology
and medicine
Nervous system of C. elegans
worm illuminated by GFP
GFP fused to Huntingtin protein
in monkey to study
Huntington’s disease
2. bla: β-lactamase enzyme can destroy
penicillin and other β-lactam antibiotics
Antibiotic resistance genes are found commonly
on plasmids and can be shared between
bacteria by conjugation
Bacterial conjugation is sex
without reproduction
Genes for making a sex pilus also are often
found encoded on plasmids
3. araC: araC regulates pGLO expression
through the presence of arabinose
Arabinose is a 5-carbon sugar, different from ribose
Gene Regulation
Q: What is “regulation”?*
"When I was warning about the danger ahead on Wall Street months ago because of the
lack of oversight, Senator McCain was telling the Wall Street Journal -- and I quote -- 'I'm
always for less regulation.' " – Sen. Barack Obama
“Senator Obama was silent on the regulation of Fannie Mae and Freddie Mac, and his
Democratic allies in Congress opposed every effort to rein them in…last year he said that
subprime loans had been, quote, “a good idea.””- Sen. John McCain
* Slide created, September 2008
Regulation means control
Prokaryotic cells require
efficiency
Eukaryotic cells must
differentiate
Eukaryotic genes can be turned on and
off
• Females only use a single X chromosome per cell
• Genes from the other chromosome are not used in that
cell
Random inactivation of one X chromosome
creates a tortoiseshell pattern in cat fur
Genes are regulated in eukaryotes in
more complex ways
• Each step in the process
of gene expression is a
possible point of control
• The cell capitalizes on
each one
The job of master control genes is to
turn many genes on or off
The gene eyeless turns on many
genes involved in formation of eyes
When eyeless is mutated, eyes
do not form
Master control genes control
formation of entire organs
-Drosophila eyeless gene can be artificially turned on in non-eye
cells
Prokaryotes must also regulate genes
Example: β- Galactosidase can hydrolyze lactose
H 2O
galactose
lactose
b-galactosidase
(aka lactase in humans)
glucose
11
b-galactosidase
10
When lactose is present, transcription
is activated
In the absence of lactose, the lac operon is
repressed by the lac repressor protein
araC allows
expression of
arabinosedigesting genes
in the presence
of arabinose
5/23/2017
73
In pGLO, arabinose-digesting genes are removed, and
araC is fused to GFP
• How do you think this
fusion was made?
• What are the structural
sequences? The
regulatory sequences?
• What happens when
we add arabinose sugar
to these bacteria?
• What do you think is
meant by “reporter
gene”?
On pGLO, the regulatory regions of the
Arabinose operon have been glued to the
structural sequences for GFP
ara GFP Operon
ara Operon
B
ara
C
A D
araC
GFP Gene
Effector (Arabinose)
Effector (Arabinose)
Gene Regulation
B A D
araC
araC
GFP Gene
RNA Polymerase
RNA Polymerase
araC
B A D
araC
GFP Gene
What will happen on the Ara (+) plates?
What will happen on the Ara (-) plates?
Grow? Glow?
Which colonies will glow?



Follow protocol
On which plates will
colonies grow?
Which colonies will
glow?
5/23/2017
76
Plasmids with novel structural and regulatory genes are
now easily manipulated by undergraduates
iGEM is an annual undergrad
bioengineering contest
Engineering bacteria to smell
better
Which strand carries the DNA's instructions for
synthesizing a particular protein from the nucleus to
the cytoplasm?
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A) Transfer RNA
B) Messenger RNA
C) RNA transcriptase
D) Ribosomal RNA
Why are the complementary base pairing rules so
important when a cell needs to copy its DNA?
• A) The cells need both strands to be accurate
because each strand codes for one half of the gene.
• B) As long as there is one strand, a copy can be made
by following the pairing rules.
• C) Unless the bases pair up correctly, the DNA strand
can break apart.
• D) Transcription cannot proceed unless the pairs are
in the proper sequence.