HUMAN GENOME PROJECT HUMAN GENOME PROJECT What is the Human Genome Project? What is the Human Genome Project? Goal: Sequence all of the nucleotides in the human DNA sequence (“genome”).
HUMAN GENOME PROJECT
Jan 25, 2016
HUMAN GENOME PROJECT. What is the Human Genome Project?. Goal: Sequence all of the nucleotides in the human DNA sequence (“genome”). HUMAN GENOME PROJECT. Why:. A. To understand how genes work. B. To understand why some genes don’t work. HUMAN GENOME PROJECT. Who:. - PowerPoint PPT Presentation
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HUMAN GENOME PROJECTHUMAN GENOME PROJECT
What is the Human Genome Project?What is the Human Genome Project?
Goal: Sequence all of the nucleotides in the human DNA sequence (“genome”).
HUMAN GENOME PROJECTHUMAN GENOME PROJECT
B. To understand why some genes don’t work.
Why: A. To understand how genes work.
B. International Project
Who: A. National Institutes of Health, Dept. of Energy
When: A. 1990; finish in 15 years
HUMAN GENOME PROJECTHUMAN GENOME PROJECT
““Cracking the Cracking the Code of Life” Code of Life” Chapter 7 first Chapter 7 first segmentsegment
When: B. First chromosome sequenced (22) - 1996
HUMAN GENOME PROJECTHUMAN GENOME PROJECT
C. 1/3 of genome completed 1999
NOTE – all Cracking the Code segments can be found atNOTE – all Cracking the Code segments can be found at
http://www.pbs.org/wgbh/nova/genome/program.htmlhttp://www.pbs.org/wgbh/nova/genome/program.html
CeleraCelera
““Cracking the Code of Cracking the Code of Life” Chapter 4 (6:29)Life” Chapter 4 (6:29)
A. Private company founded by Craig Venter, former NIH scientist
B. Finish project in 2 years?
How do you profit from sequencing How do you profit from sequencing the human genome?the human genome?
““Cracking the Code of Life” Chapter 8 (4:06)Cracking the Code of Life” Chapter 8 (4:06)
Sell information for scientists to look at.Sell information for scientists to look at.
Eventually, public project will complete HGP, so Eventually, public project will complete HGP, so what do you sell then?what do you sell then?
““Cracking the Code of Life” Chapter 7 second Cracking the Code of Life” Chapter 7 second segment (49:10).segment (49:10).
How do you profit? continuedHow do you profit? continued
Patenting DNA sequences – is this right?Patenting DNA sequences – is this right?
Whose data is it?Whose data is it?
Does patenting DNA sequences Does patenting DNA sequences encourage or discourage research from encourage or discourage research from being done?being done?
Who won?Who won?
Both groups shared credit for “finishing” Both groups shared credit for “finishing” the HGP in 2001. the HGP in 2001.
Competition sped up sequencing process.Competition sped up sequencing process.
““Cracking the Code of Life” link?Cracking the Code of Life” link?
What Have We Learned From What Have We Learned From HGP?HGP?
Humans are 99.9% identical.Humans are 99.9% identical.
Total number of genes ~ 30,000. This Total number of genes ~ 30,000. This doesn’t match the number of proteins doesn’t match the number of proteins (over 100,000) so each gene must be able (over 100,000) so each gene must be able to code for more than one protein.to code for more than one protein.
Over 50% of genes have unknown Over 50% of genes have unknown functions.functions.
What Have We Learned From What Have We Learned From HGP?HGP?
Less than 2% of DNA codes for genes.Less than 2% of DNA codes for genes.
Most genes are clustered in “urban Most genes are clustered in “urban centers” (not randomly spread out).centers” (not randomly spread out).
Over 50% of DNA is “not human” – Over 50% of DNA is “not human” – hitchhiking “junk” DNA.hitchhiking “junk” DNA.
What’s next?What’s next?
Gene regulation – how do genes know Gene regulation – how do genes know when to turn on and off?when to turn on and off?
Proteome – what proteins do these genes Proteome – what proteins do these genes code for and what do the proteins do?code for and what do the proteins do?
Personalized medicine – medications to Personalized medicine – medications to treat you based on your genetics.treat you based on your genetics.
What’s next?What’s next?
Copy Number Variant – readingCopy Number Variant – reading
SNP’s – readingSNP’s – reading
Epigenetics – readingEpigenetics – reading
STR’s - labSTR’s - lab
How does sequencing work?How does sequencing work?
The Key
This is a nucleotide called a
dideoxynucleotide.
Missing oxygen = deoxyribonucleic acid
Missing oxygen #2= dideoxyribonucleic acid
Why are dideoxynucleotides important?
Since there is no oxygen on the 3’ end, no additional nucleotides can be added.
DNA Synthesis is stopped.
What is needed for a Sequencing What is needed for a Sequencing Reaction?Reaction?
Original DNAOriginal DNA
NucleotidesNucleotides
PrimerPrimer
DNA PolymeraseDNA Polymerase
““Detectable” dideoxynucleotides Detectable” dideoxynucleotides (radioactivity or fluorescence)(radioactivity or fluorescence)
Now it’s your turn to sequence!Now it’s your turn to sequence!
How does a Sequencing Reaction How does a Sequencing Reaction work?work?
www.dnai.orgwww.dnai.org- manipulation- manipulation
- techniques- techniques- sorting and sequencing- sorting and sequencing- cycle sequencing- cycle sequencing
Three stepsThree steps1. Denaturing – 951. Denaturing – 9500CC2. Annealing – 502. Annealing – 5000CC3. Extension – 603. Extension – 6000CC
Only one cycle so do not need to use expensive Taq polymerase
How does a Sequencing Reaction How does a Sequencing Reaction work?work?
Nucleotides are randomly selected by DNA Nucleotides are randomly selected by DNA Polymerase.Polymerase.
Sequencing is stopped when ddNTP is randomly Sequencing is stopped when ddNTP is randomly selected. selected.
Sequences of varying lengths are produced.Sequences of varying lengths are produced.
How would we separate these differently sized How would we separate these differently sized pieces?pieces?
How does a Sequencing Reaction How does a Sequencing Reaction work?work?
Gel ElectrophoresisGel Electrophoresis
Laser detects the Laser detects the fluorescence of each fluorescence of each ddNTPddNTP
Computer records the Computer records the order of the colors order of the colors (order of the bases)(order of the bases)
How does a Sequencing Reaction How does a Sequencing Reaction work?work?
Results are presented as Results are presented as an “electropherogram”.an “electropherogram”.
www.dnai.orgwww.dnai.org
- manipulation- manipulation
- techniques- techniques
- Interview- Interview
““Inside an automated Inside an automated sequencer”.sequencer”.
Sequencing Process ReviewSequencing Process Review
Sequencing AnimationSequencing Animation
Now it’s your turn to sequence, Now it’s your turn to sequence, Part 2!Part 2!
How do you sequence so many How do you sequence so many letters so quickly?letters so quickly?
Shotgun sequencingShotgun sequencing – – divide many copies of divide many copies of genome into small genome into small bits. Sequence each bits. Sequence each fragment. Use fragment. Use computers to align computers to align sequence.sequence.
How do you sequence so many How do you sequence so many letters so quickly?letters so quickly?
www.dnai.org www.dnai.org
- genome- genome
- The Project- The Project
- Putting It Together- Putting It Together
- Animations- Animations
- Whole Genome Shotgun (private)- Whole Genome Shotgun (private)
How do you sequence so many How do you sequence so many letters so quickly?letters so quickly?
www.dnai.org www.dnai.org
- genome- genome
- The Project- The Project
- Putting It Together- Putting It Together
- Sequencing Game- Sequencing Game
So what can you conclude about So what can you conclude about shotgun sequencing?shotgun sequencing?
Overlapping provides a Overlapping provides a contextcontext. (unlike first . (unlike first Mouse and Cookie sentence fragments).Mouse and Cookie sentence fragments).
Requires multiple copies each copy cut with a Requires multiple copies each copy cut with a different restriction enzyme to generate different restriction enzyme to generate overlapping piecesoverlapping pieces
Up to 8% of human genome remains Up to 8% of human genome remains unsequenced due to highly repetitive sections unsequenced due to highly repetitive sections (especially ends and middles– telomeres and (especially ends and middles– telomeres and centromeres).centromeres).
Public – a random couple from Buffalo, NYPublic – a random couple from Buffalo, NY
Celera – random, nameless volunteers Celera – random, nameless volunteers (though Dr. Venter’s DNA was “randomly” (though Dr. Venter’s DNA was “randomly” selectedselected
Whose DNA was sequenced?Whose DNA was sequenced?
What’s next?What’s next?
To learn which sequences lead to genetic To learn which sequences lead to genetic disorders, many different human genomes disorders, many different human genomes need to be sequenced.need to be sequenced.
Which is more important to Which is more important to studying genetic disease?studying genetic disease?
Sequences that are the Sequences that are the samesame??
Sequences that are Sequences that are differentdifferent??
WHY?
What are those differences called?What are those differences called?
SNP’sSNP’s – single nucleotide – single nucleotide polymorphisms; DNA polymorphisms; DNA sequence that is one letter sequence that is one letter different.different.
Develop “personalized Develop “personalized medicine” based on the exact medicine” based on the exact SNP causing genetic disorder.SNP causing genetic disorder.
Are SNP’s the whole story?Are SNP’s the whole story?
CNV’SCNV’S – copy number variants; not everyone – copy number variants; not everyone has two copies of each gene.has two copies of each gene.
Higher number of gene copies, higher level of Higher number of gene copies, higher level of protein might be produced; not necessarily protein might be produced; not necessarily good.good.
Ex. EGFR copy number can be higher than Ex. EGFR copy number can be higher than normal in some types of lung cancer cells.normal in some types of lung cancer cells.
Copy Number VariantsCopy Number Variants
What else is next?What else is next?
Epigenome Epigenome – changes made to DNA – changes made to DNA structure without altering the sequence of structure without altering the sequence of bases.bases.
These changes quite often involve a These changes quite often involve a methyl (-CHmethyl (-CH33) group to tag or mark a gene.) group to tag or mark a gene.
Cell normally uses these methyl tags to Cell normally uses these methyl tags to “turn off” a gene.“turn off” a gene.
DNA or histones are DNA or histones are methylated.methylated.
Does this mean that identical twins Does this mean that identical twins don’t have to be . . . Identical?don’t have to be . . . Identical?
YES! Think of YES! Think of the Agouti the Agouti mice.mice.
Only difference Only difference is what the is what the mom ate prior mom ate prior to conception to conception and birth.and birth.
Other epigenome examples?Other epigenome examples?
Let’s go to the video!Let’s go to the video!
So what’s the sequencing So what’s the sequencing “revolution”?“revolution”?
Original sequencing Original sequencing reactions used reactions used radioactive ddNTP’s radioactive ddNTP’s not fluorescent.not fluorescent.
Results looked like:Results looked like:
Problems with Radioactive Problems with Radioactive SequencingSequencing
Very difficult to read Very difficult to read resultsresults
Cannot reuse a Cannot reuse a machine exposed to machine exposed to radioactivity in a radioactivity in a machinemachine
Again, what’s the “revolution”?Again, what’s the “revolution”?
Computers and fluorescent ddNTP’sComputers and fluorescent ddNTP’s
Machines can automatically run a sequencing Machines can automatically run a sequencing reaction.reaction.
Computers can store sequencing data.Computers can store sequencing data.
Fluroescent ddNTP’s make machines reusable.Fluroescent ddNTP’s make machines reusable.
““100 letters in a day vs. 1000 letters every 100 letters in a day vs. 1000 letters every second”second”
More HGP InfoMore HGP Info
Cracking the Code of Life – Chapters 4, 5, 6, Cracking the Code of Life – Chapters 4, 5, 6, and 16 and 16 (http://www.pbs.org/wgbh/nova/genome/program(http://www.pbs.org/wgbh/nova/genome/program.html).html)
In order to sequence In order to sequence allall DNA, Celera relied on DNA, Celera relied on freely available DNA sequence from public freely available DNA sequence from public research group.research group.
Who finished first – public or private research Who finished first – public or private research group? When?group? When?
ELSI?ELSI?
Ethical, Legal, and Social Issues Ethical, Legal, and Social Issues
At the beginning of the project, genetic privacy At the beginning of the project, genetic privacy was one of the major concerns – as we learn was one of the major concerns – as we learn more about our own DNA sequences:more about our own DNA sequences:
– Who should have access to that information?Who should have access to that information?
– How do you help someone interpret that information How do you help someone interpret that information and decide how to act on it?and decide how to act on it?
ELSI?ELSI?
ELSI Video
Human Genome ProjectHuman Genome Project
Cracking the Code of Life – Chapter Two Cracking the Code of Life – Chapter Two “Getting the Letters Out”“Getting the Letters Out”
http://www.pbs.org/wgbh/nova/genome/http://www.pbs.org/wgbh/nova/genome/program.htmlprogram.html
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