Current sequencing technology and Human Genome Project

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Current sequencing technologyand Human Genome
Project

• Original Presenter
• Under guidance of Dr. Jae K. Lee,
• October 3, 2000
• -Nitin Jain
• Cited by David Fu
• 2002/4/23

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Content

• Importance of DNA sequencing
• Modern methods of sequencing
• Chemical method of sequencing
• Enzymatic method of sequencing
• Automated sequencing
• Human Genome project (HGP)
• Interesting facts about HGP
• Future goals
• Advantages of HGP

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Importance of DNA sequencing

• Confirmation of the right fragment
• Know the restiction sites in the fragment
• Check the quality of DNA
• Modification of Genes

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Methods of DNA Sequencing

• Maxam Gilbert's method
• Fred Sanger's method
• Automated sequencing method
• DNA-chip sequencing (under development)
• Sequencing of individual bases (at the atomic
  level)

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Maxam Gilbert method(Chemical Method)

• Radioactive labeling of 5end
• Denaturing the double stranded DNA
• Random degradation of specific nucleotides
• Polyacrylamide Gel separation
• Autoradiography
• Four parallel processes

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Chemical Method of Sequencing
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Sangers method(Enzyatic method)

• Understand the DNA synthesis (PCR) first
• Template
• Primer
• Deoxy-ribonucleoside tri-phosphates (dNTPs)
• Enzyme
• Reaction Buffer

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Di-deoxynucleotides (ddNTPs)
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Sangers method

• Uses small amount of di-deoxy-ribonucleoside
  triphosphates (ddNTPs) in conjunction with dNTPs
• Random incorporation of ddNTPs inhibits reaction
• Ladder of fragments detected by label (chemical
  or radioactive)
• Four parallel processes

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Automated sequencing

• Modification of Sangers method
• Fluorescently labelled di-deoxy bases are used,
  allowing all four nucleotides to be put in the
  same lane
• Laser detector, different excitation frequencies
  of differencet labels
• Diffenernt wavelengths produce different peaks of
  different colours.

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Automatic sequencing
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Human Genome Project (HGP)

• Refers to the euchromatic portion of the human
  genome
• Size of human genome 3 billion bases
• Total number of Genes 100,000

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Some interesting facts of HGP

• International consortium of scientists from 16
  institutes around the world
• Total cost over 15 year period 3 billion
• An initial "working draft", which covers gt90 of
  the genome to a depth of 3-4x redundant sequence,
  is over
• Average proportion of G-C base pairs across
  genome 41

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Advantages of HGP

• Creating the reference sequence or blueprint
  of humans
• Comprehensive catalogue of the Single Nucleotide
  Polymorphisms (SNPs)
• Understanding the diseases and cure them
• Revolutionize medicine
• Create healthier (super) humans