DNA, Genes, and Biotechnology Chapter 22. Structure of the Hereditary Material Experiments in the...
Transcript of DNA, Genes, and Biotechnology Chapter 22. Structure of the Hereditary Material Experiments in the...
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DNA, Genes, and Biotechnology
Chapter 22
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Structure of the Hereditary Material
• Experiments in the 1950s showed that DNA is the hereditary material
• Scientists raced to determine the structure of DNA
• 1953 - Watson and Crick proposed that DNA is a double helix
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Structure of Nucleotides in DNA
• Each nucleotide consists of
– Deoxyribose (5-carbon sugar)
– Phosphate group
– A nitrogen-containing base
• Four bases
– Adenine, Guanine, Thymine, Cytosine(A) (G) (T) (C)
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Nucleotide Bases
phosphate group
deoxyribose
ADENINE (A)
THYMINE (T)
CYTOSINE (C)
GUANINE (G)
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Watson-Crick Model
• DNA consists of two nucleotide strands
• Strands run in opposite directions
• Strands are held together by hydrogen
bonds between bases
• A binds with T, C with G
• Molecule is a double helix
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Structure of DNA
A gene is a sequence of nucleotides in a DNA molecule
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DNA Structure Helps Explain How It Duplicates
• DNA is two nucleotide strands held
together by hydrogen bonds
• Hydrogen bonds between two strands
are easily broken
• Each single strand then serves as
template for new strand
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DNA Replication
newnew old old
• Each parent strand
remains intact
• Every DNA
molecule is half
“old” and half “new”
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Base-Pairing during
Replication
Each old strand serves as the template for complementary new strand
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Errors in DNA Replication
• Mistakes can occur during replication
• Enzymes may detect and correct the
problem, restoring the proper DNA
sequence
• When the error is not corrected, the
result is a mutation
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Gene Mutations
Base-Pair Substitutions
Insertions
Deletions
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Effect of Base-Pair Substitution
original base triplet in a DNA strand
As DNA is replicated, proofreadingenzymes detect the mistake andmake a substitution for it:
a base substitution within the triplet (red)
One DNA molecule carries the original, unmutated sequence
The other DNAmolecule carries a gene mutation
POSSIBLE OUTCOMES:
OR
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Frameshift Mutations
• Insertion
– Extra base added into gene region
• Deletion
– Base removed from gene region
• Both shift the reading frame
• Result in many wrong amino acids
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Frameshift Mutation
ARGININE GLYCINE TYROSINE TRYPTOPHAN ASPARAGINE
ARGININE GLYCINE LEUCINE GLUTAMATELEUCINE
mRNA
PARENTAL DNA
amino acid sequence
altered mRNA
BASE INSERTION
altered amino-acid sequence
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Transposable Elements
• DNA segments that move
spontaneously about the genome
• When they insert into a gene
region, they usually inactivate
that gene
• Neurofibromatosis
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DNA to RNA to Proteins
transcriptionDNA RNA
translationprotein
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Three Classes of RNAs
• Messenger RNA
– Carries protein-building instruction
• Ribosomal RNA
– Major component of ribosomes
• Transfer RNA
– Delivers amino acids to ribosomes
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Base-Pairing during Transcription
• A new RNA strand can be put together
on a DNA region
DNA
DNA DNA
RNAG C A T
C G T A
G C A U
C G T A
base-pairing in DNA replication base-pairing in transcription
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Promoter
• A base sequence in the DNA that signals the start of a gene
• For transcription to occur, RNA polymerase must first bind to a promoter
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Gene Transcription
transcribed DNA winds up again
DNA to be transcribed unwinds
mRNAtranscript
RNA polymerase
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Transcript Modificationunit of transcription in a DNA strand
exon intron
mature mRNA transcript
poly-A tail
5’
5’ 3’
3’
snipped out
snipped out
exon exonintron
cap
transcription into pre-mRNA
3’ 5’
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Gene Regulation
• Most cells of your body carry the same
genes
• Each uses only a tiny subset at any
time, and some are never turned on
• Regulatory proteins can speed up or
halt transcription
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Genetic Code
• Set of 64 base triplets
• Codons– Nucleotide bases read in
blocks of three
• 61 specify amino acids
• 3 stop translation
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Code Is Redundant
• Twenty kinds of amino acids are specified by
61 codons
• Most amino acids can be specified by more
than one codon
• Six codons specify leucine
– UUA, UUG, CUU, CUC, CUA, CUG
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tRNA Structure
codon in mRNA
anticodon in tRNA
amino acid OH
tRNA molecule’s attachment site for amino acid
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Ribosomestunnel
small ribosomal subunit large ribosomal subunit intact ribosome
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Three Stages of Translation
Initiation
Elongation
Termination
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Initiation
• Initiator tRNA binds to small ribosomal subunit
• Small subunit/tRNA complex attaches to mRNA and moves along it to an AUG “start” codon
• Large ribosomal subunit joins complex
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Binding Sites on Large Subunit
binding site for mRNA
P (first binding site for tRNA)
A (second binding site for tRNA)
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Elongation
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Termination
• A stop codon in the mRNA moves onto the ribosomal binding site
• No tRNA has a corresponding anticodon
• Proteins called release factors bind to the ribosome
• mRNA and polypeptide are released
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Making Recombinant DNA
5’
3’
G
C T T A A
A A T T C
G
G A A T T C
C T T A A G3’
5’
one DNA fragment another DNA fragment
3’
5’
Restriction enzymes cut DNA into fragments
Fragments base-pair at sticky ends
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Using Plasmids
• Plasmid is small circle of bacterial DNA
• Foreign DNA can be inserted into
plasmid
– Forms recombinant plasmids
– Plasmid is a cloning vector
– Can be used to deliver DNA into
another cell
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Using Plasmids
DNA fragments+enzymes
recombinantplasmids
host cells containing recombinant plasmids
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Polymerase Chain Reaction
• Sequence to be copied is heated
• Primers are added and bind to ends of single strands
• DNA polymerase uses free nucleotides to create complementary strands
• Doubles number of copies of DNA
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PCR (1)
double-stranded DNA to copy
DNA heated to 90°– 94°C
primers added to base-pair with ends
mixture cooled; base-pairing of primers and ends of DNA strands
DNA polymerasesassemble new DNA strands
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PCR (2)mixture heated again; makes all DNA fragments unwind
mixture cooled; base-pairing between primers and ends of single DNA strands
DNA polymerase action again doubles number of identical DNA fragments
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DNA Sequencing:Reaction Mixture
• Copies of DNA to be sequenced
• Primer
• DNA polymerase
• Standard nucleotides
• Modified nucleotides
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Reactions Proceed
• Nucleotides are assembled to create complementary strands
• When a modified nucleotide is included, synthesis stops
• Result is millions of tagged copies of varying length
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Recording the Sequence
T C C A T G G A C CT C C A T G G A C
T C C A T G G A
T C C A T G G
T C C A T G
T C C A T
T C C A
T C C
T C
T
electrophoresisgel
one of the many fragments of DNA migratingthrough the gel
one of the DNA fragmentspassing through a laser beam after moving through the gel
T C C A T G G A C C A
•DNA is placed on gel
•Fragments move off
gel in size order; pass
through laser beam
•Color each fragment
fluoresces is recorded
on printout
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The Human Genome Project
Goal - Map the entire human genome
• Initially thought by many to be a waste of resources
• Process accelerated when Craig Ventner used bits of cDNAs as hooks to find genes
• Sequencing was completed ahead of schedule in early 2001
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Surprising Discoveries
• Coding regions (exons) make up only 1.5% of our DNA
• About half of remaining DNA is repeated segments
• Around 1.4 million SNPs in the human genome (single nucleotide polymorphisms)
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Using Human Genes
• Even with gene in hand it is difficult to manipulate it to advantage
• Viruses usually used to insert genes into cultured human cells but procedure has problems
• Very difficult to get modified genes to work where they should
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DNA Fingerprints
• Unique array of DNA fragments
• Inherited from parents in Mendelian
fashion
• Even full siblings can be distinguished
from one another by this technique
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Tandem Repeats
• Short regions of DNA that differ substantially among people
• Many sites in genome where tandem repeats occur
• Each person carries a unique combination of repeat numbers
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RFLPs
• Restriction fragment-length polymorphisms
• DNA from areas with tandem repeats is cut with restriction enzymes
• Because of the variation in the amount of repeated DNA, the restriction fragments vary in size
• Variation is detected by gel electrophoresis
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Biotechnology Concerns
• Mutation of transgenic bacteria or
viruses could yield new pathogens
• Bioengineered plants and animals could
alter ecological balance
• Genetic screening could lead to
discrimination
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Engineered Bacteria
• Transgenic bacteria can be used to
grow medically valuable proteins
– Insulin, interferon, blood-clotting factors
– Vaccines
• Human gene is inserted into bacteria,
which are then grown in huge vats
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Engineered Plants and Animals
• Transgenic goats produce human
tissue plasminogen activator (tPA)
• Chinese hamster ovary cells produce
blood-clotting factor VIII
• Aspen plants produce less lignin and
more cellulose
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Jefferson’s Genes
• It was suspected that Jefferson had at least one child with his slave, Sally Hemings
• DNA evidence showed that a modern male descendant of Sally Hemings has a Y chromosome that had been passed down to him through the generations from Thomas Jefferson