HUMAN GENOME PROJECT

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HUMAN GENOME PROJECT

• What is the Human Genome Project?

Goal Sequence all of the nucleotides in the
human DNA sequence (genome).
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HUMAN GENOME PROJECT
Why
A. To understand how genes work.
B. To understand why some genes dont work.
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HUMAN GENOME PROJECT
Who
A. National Institutes of Health, Dept. of Energy
B. International Project
When
A. 1990 finish in 15 years
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HUMAN GENOME PROJECT
When
B. First chromosome sequenced (22) - 1996
C. 1/3 of genome completed 1999

• Cracking the Code of Life Chapter 7 first
  segment

• NOTE all Cracking the Code segments can be
  found at
• http//www.pbs.org/wgbh/nova/genome/program.html

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Celera
A. Private company founded by Craig Venter,
former NIH scientist
B. Finish project in 2 years?

• Cracking the Code of Life Chapter 4 (629)

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How do you profit from sequencing the human
genome?

• Cracking the Code of Life Chapter 8 (406)
• Sell information for scientists to look at.
• Eventually, public project will complete HGP, so
  what do you sell then?
• Cracking the Code of Life Chapter 7 second
  segment (4910).

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How do you profit? continued

• Patenting DNA sequences is this right?
• Whose data is it?
• Does patenting DNA sequences encourage or
  discourage research from being done?

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Who won?

• Both groups shared credit for finishing the HGP
  in 2001.
• Competition sped up sequencing process.
• Cracking the Code of Life link?

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What Have We Learned From HGP?

• Humans are 99.9 identical.
• Total number of genes 30,000. This doesnt
  match the number of proteins (over 100,000) so
  each gene must be able to code for more than one
  protein.
• Over 50 of genes have unknown functions.

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What Have We Learned From HGP?

• Less than 2 of DNA codes for genes.
• Most genes are clustered in urban centers (not
  randomly spread out).
• Over 50 of DNA is not human hitchhiking
  junk DNA.

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Whats next?

• Gene regulation how do genes know when to turn
  on and off?
• Proteome what proteins do these genes code for
  and what do the proteins do?
• Personalized medicine medications to treat you
  based on your genetics.

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Whats next?

• Copy Number Variant reading
• SNPs reading
• Epigenetics reading
• STRs - lab

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How does sequencing work?
The Key
Missing oxygen deoxyribonucleic acid
Missing oxygen 2 dideoxyribonucleic acid
nucleotide called a
This is a
dideoxynucleotide.
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Why are dideoxynucleotides important?

• Since there is no oxygen on the 3 end, no
  additional nucleotides can be added.
• DNA Synthesis is stopped.

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What is needed for a Sequencing Reaction?

• Original DNA
• Nucleotides
• Primer
• DNA Polymerase
• Detectable dideoxynucleotides (radioactivity or
  fluorescence)

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Now its your turn to sequence!
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How does a Sequencing Reaction work?

• www.dnai.org
• - manipulation
• - techniques
• - sorting and sequencing
• - cycle sequencing
• Three steps
• 1. Denaturing 950C
• 2. Annealing 500C
• 3. Extension 600C

Only one cycle so do not need to use expensive
Taq polymerase
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How does a Sequencing Reaction work?

• Nucleotides are randomly selected by DNA
  Polymerase.
• Sequencing is stopped when ddNTP is randomly
  selected.
• Sequences of varying lengths are produced.
• How would we separate these differently sized
  pieces?

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How does a Sequencing Reaction work?

• Gel Electrophoresis
• Laser detects the fluorescence of each ddNTP
• Computer records the order of the colors (order
  of the bases)

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How does a Sequencing Reaction work?

• Results are presented as an electropherogram.
• www.dnai.org
• - manipulation
• - techniques
• - Interview
• Inside an automated sequencer.

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Sequencing Process Review

• Sequencing Animation

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Now its your turn to sequence, Part 2!
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How do you sequence so many letters so quickly?

• Shotgun sequencing divide many copies of genome
  into small bits. Sequence each fragment. Use
  computers to align sequence.

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How do you sequence so many letters so quickly?

• www.dnai.org
• - genome
• - The Project
• - Putting It Together
• - Animations
• - Whole Genome Shotgun (private)

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How do you sequence so many letters so quickly?

• www.dnai.org
• - genome
• - The Project
• - Putting It Together
• - Sequencing Game

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So what can you conclude about shotgun sequencing?

• Overlapping provides a context. (unlike first
  Mouse and Cookie sentence fragments).
• Requires multiple copies each copy cut with a
  different restriction enzyme to generate
  overlapping pieces
• Up to 8 of human genome remains unsequenced due
  to highly repetitive sections (especially ends
  and middles telomeres and centromeres).

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Whose DNA was sequenced?

• Public a random couple from Buffalo, NY
• Celera random, nameless volunteers (though Dr.
  Venters DNA was randomly selected

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Whats next?

• To learn which sequences lead to genetic
  disorders, many different human genomes need to
  be sequenced.

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Which is more important to studying genetic
disease?

• Sequences that are the same?
• Sequences that are different?

WHY?
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What are those differences called?

• SNPs single nucleotide polymorphisms DNA
  sequence that is one letter different.
• Develop personalized medicine based on the
  exact SNP causing genetic disorder.

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Are SNPs the whole story?

• CNVS copy number variants not everyone has
  two copies of each gene.
• Higher number of gene copies, higher level of
  protein might be produced not necessarily good.
• Ex. EGFR copy number can be higher than normal in
  some types of lung cancer cells.

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Copy Number Variants
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What else is next?

• Epigenome changes made to DNA structure without
  altering the sequence of bases.
• These changes quite often involve a methyl (-CH3)
  group to tag or mark a gene.
• Cell normally uses these methyl tags to turn
  off a gene.

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• DNA or histones are methylated.

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Does this mean that identical twins dont have to
be . . . Identical?

• YES! Think of the Agouti mice.
• Only difference is what the mom ate prior to
  conception and birth.

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Other epigenome examples?

• Lets go to the video!

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So whats the sequencing revolution?

• Original sequencing reactions used radioactive
  ddNTPs not fluorescent.
• Results looked like

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Problems with Radioactive Sequencing

• Very difficult to read results
• Cannot reuse a machine exposed to radioactivity
  in a machine

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Again, whats the revolution?

• Computers and fluorescent ddNTPs
• Machines can automatically run a sequencing
  reaction.
• Computers can store sequencing data.
• Fluroescent ddNTPs make machines reusable.
• 100 letters in a day vs. 1000 letters every
  second

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More HGP Info

• Cracking the Code of Life Chapters 4, 5, 6, and
  16 (http//www.pbs.org/wgbh/nova/genome/program.ht
  ml)
• In order to sequence all DNA, Celera relied on
  freely available DNA sequence from public
  research group.
• Who finished first public or private research
  group? When?

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ELSI?

• Ethical, Legal, and Social Issues
• At the beginning of the project, genetic privacy
  was one of the major concerns as we learn more
  about our own DNA sequences
• Who should have access to that information?
• How do you help someone interpret that
  information and decide how to act on it?

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ELSI?
ELSI Video
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Human Genome Project

• Cracking the Code of Life Chapter Two Getting
  the Letters Out
• http//www.pbs.org/wgbh/nova/genome/program.html