A Lot More Advanced Biotechnology Tools

Slides:



Advertisements
Similar presentations
Human Genome Project What did they do? Why did they do it? What will it mean for humankind? Animation OverviewAnimation Overview - Click.
Advertisements

Basic Molecular Biology Many slides by Omkar Deshpande.
Intro to DNA Sequencing
A Lot More Advanced Biotechnology Tools DNA Sequencing.
DNA sequencing Matt Hudson. DNA Sequencing Dideoxy sequencing was developed by Fred Sanger at Cambridge in the 1970s. Often called “Sanger sequencing”.
DNA Sequencing. DNA sequencing … ACGTGACTGAGGACCGTG CGACTGAGACTGACTGGGT CTAGCTAGACTACGTTTTA TATATATATACGTCGTCGT ACTGATGACTAGATTACAG ACTGATTTAGATACCTGAC.
DNA Sequencing How do you do it?. DNA Sequencing DNA sequencing – used to determine the actual DNA sequence of an organism. Using a computer, one can.
Genome sequencing MUPGRET Workshop Joe Polacco. Size of human genome 23 pairs of chromosomes 3.1 billion bp If code written in NYC phone books and stacked.
Goals of the Human Genome Project determine the entire sequence of human DNA identify all the genes in human DNA store this information in databases improve.
Introduction to DNA Sequencing Technology. Dideoxy Sequencing (Sanger Sequencing, Chain Terminator method). Clone the fragments to be sequenced into the.
7.1 cont’d: Sanger Sequencing SBI4UP MRS. FRANKLIN.
DNA Sequencing Today, laboratories routinely sequence the order of nucleotides in DNA. DNA sequencing is done to: Confirm the identity of genes isolated.
Recombinant DNA Technology for the non- science major.
Manipulating DNA Genetic Engineering uses the understanding of the properties of DNA to study and change DNA sequences in living organisms – Invitro… in.
Let’s Review Biotechnology! (Why not?). Embryonic Cloning Label parts and steps. + ?
Electrophoresis & RFLPs
Chapter 14 Genomes and Genomics. Sequencing DNA dideoxy (Sanger) method ddGTP ddATP ddTTP ddCTP 5’TAATGTACG TAATGTAC TAATGTA TAATGT TAATG TAAT TAA TA.
AP Biology A Lot More Advanced Biotechnology Tools Sequencing.
Applications of DNA technology
Section 2 Genetics and Biotechnology DNA Technology
Genomics & Proteomics Analysis Chapter 20 Overview of topics to be discussed  How to sequence genomic DNA (we will have to touch briefly on polymerase.
A Lot More Advanced Biotechnology Tools (Part 1) Sequencing.
A Sequenciação em Análises Clínicas Polymerase Chain Reaction.
Genetics 7: Analyzing DNA Sequences DNA Sequencing Determining base by base the nucleotide sequence of a fragment of DNA.
Restriction Fragments and Mapping Restriction Fragment Analysis – System used to compare the genes and DNA sequences between individuals in a population.
+ DNA Technology 1. + DNA Extraction Chemical treatments Chemical treatments cause cells and nuclei to burst sticky The DNA is inherently sticky, and.
Genome Characterization DNA sequence-ULTIMATE Map DNA sequencing-methods Assembly/sequencing BIO520 BioinformaticsJim Lund Assigned reading: Service 2006.
GENE SEQUENCING. INTRODUCTION CELL The cells contain the nucleus. The chromosomes are present within the nucleus.
© Copyright 2010 Robert D. Conway All Rights Reserved Who Invented It? The Controversial History of Technology and Invention
DNA Sequencing.
A Lot More Advanced Biotechnology Tools Sequencing.
Sequencing by the Sanger Dideoxynucleotide Chain Termination Method 1. Prepare replication template denature, add synthetic primer, promote annealing TAGGCGA.
Johnson - The Living World: 3rd Ed. - All Rights Reserved - McGraw Hill Companies Genomics Chapter 10 Copyright © McGraw-Hill Companies Permission required.
Genome They are the volums of an encyclopaedia called Genome. Cell Nucleus Tissues The chromosomes contains the instruction of alive beings.
A Lot More Advanced Biotechnology Tools (Part 2) Sequencing.
Genomics Part 1. Human Genome Project  G oal is to identify the DNA sequence of every gene in humans Genome  all the DNA in one cell of an organism.
DNA Sequencing Mimi Chen & Joanne Kim
Chapter 17 – 18 Biotechnology: Genomics & DNA Technology.
Title: Studying whole genomes Homework: learning package 14 for Thursday 21 June 2016.
핵산 염기서열 분석(DNA SEQUENCING)
Topic Cloning and analyzing oxalate degrading enzymes to see if they dissolve kidney stones with Dr. VanWert.
Cse587A/Bio 5747: L2 1/19/06 1 DNA sequencing: Basic idea Background: test tube DNA synthesis DNA polymerase (a natural enzyme) extends 2-stranded DNA.
DNA Technology.
Restriction Fragments and Mapping
Genomics Sequencing genomes.
DNA Sequencing Techniques
Genomic Analysis Chapter 19
Di-deoxynucleotide Chain Termination
Genetic Research and Biotechnology
Section 2 Genetics and Biotechnology DNA Technology
DNA Technology.
Sequencing Technologies
DNA Technology.
The Human Genome Project
DNA Sequence Determination (Sanger)
Screening a Library for Clones Carrying a Gene of Interest
DNA Sequencing The DNA from the genome is chopped into bits- whole chromosomes are too large to deal with, so the DNA is broken into manageably-sized overlapping.
UD. IV. GENÈTICA. Ll. IV. 5. Biotecnologia
Genomes and Their Evolution
Genomic Analysis Chapter 19-20
DNA and the Genome Key Area 8a Genomic Sequencing.
A Sequenciação em Análises Clínicas
DNA Technology.
Basic Molecular Biology
Plant Biotechnology Lecture 2
DNA Sequencing.
DNA Technology.
DNA Technology.
DNA Technology.
SBI4U0 Biotechnology.
Presentation transcript:

A Lot More Advanced Biotechnology Tools Sequencing 2007-2008

DNA Sequencing Sanger method determine the base sequence of DNA based on replication dideoxynucleotides ddATP, ddGTP, ddTTP, ddCTP missing O for bonding of next nucleotide terminates the growing chain

DNA Sequencing Sanger method 1 Sanger method synthesize complementary DNA strand in vitro in each tube: “normal” N-bases dideoxy N-bases ddA, ddC, ddG, ddT DNA polymerase primer buffers & salt 2 3 4 2

Reading the sequence Load gel with sequences from ddA, ddT, ddC, ddG in separate lanes read lanes manually & carefully polyacrylamide gel

Fred Sanger 1978 | 1980 This was his 2nd Nobel Prize!! 1st was in 1958 for the structure of insulin

Advancements to sequencing Fluorescent tagging no more radioactivity all 4 bases in 1 lane each base a different color Automated reading

Advancements to sequencing Fluorescent tagging sequence data Computer read & analyzed

Advancements to sequencing Capillary tube electrophoresis no more pouring gels higher capacity & faster Applied Biosystems, Inc (ABI) built an industry on these machines 384 lanes

Big labs! economy of scale PUBLIC Joint Genome Institute (DOE) MIT Washington University of St. Louis Baylor College of Medicine Sanger Center (UK) PRIVATE Celera Genomics Celera: Rockville, MD & San Francisco, CA Baylor: Houston TX

Automated Sequencing machines Really BIG labs!

Human Genome Project U.S government project Celera Genomics begun in 1990 estimated to be a 15 year project DOE & NIH initiated by Jim Watson led by Francis Collins goal was to sequence entire human genome 3 billion base pairs Celera Genomics Craig Venter challenged gov’t would do it faster, cheaper private company 1990-1995 build the technology groundwork improve sequencing methods build clones build better data management systems (computer tools to find overlaps) better, cheaper, faster! 1996-1998 painstaking sequencing work 1998 Celera genomics challenge 2000 rough draft of human genome (90% sequence, 99% accurate) 2001 1st draft of human genome 2003 “finished” sequence of human genome can’t sequence telomeres & centromeres

Different approaches “map-based method” “shotgun method” gov’t method Craig Venter’s method 1. Cut DNA entire chromosome into small fragments and clone. 2. Sequence each segment & arrange based on overlapping nucleotide sequences. Cut DNA segment into fragments, arrange based on overlapping nucleotide sequences, and clone fragments. 2. Cut and clone into smaller fragments. 3. Assemble DNA sequence using overlapping sequences.

Human Genome Project On June 26, 2001, HGP published the “working draft” of the DNA sequence of the human genome. Historic Event! blueprint of a human the potential to change science & medicine

Sequence of 46 Human Chromosomes 3G of data 3 billion base pairs

human genome 3.2 billion bases TACGCACATTTACGTACGCGGATGCCGCGACTATGATCACATAGACATGCTGTCAGCTCTAGTAGACTAGCTGACTCGACTAGCATGATCGATCAGCTACATGCTAGCACACYCGTACATCGATCCTGACATCGACCTGCTCGTACATGCTACTAGCTACTGACTCATGATCCAGATCACTGAAACCCTAGATCGGGTACCTATTACAGTACGATCATCCGATCAGATCATGCTAGTACATCGATCGATACTGCTACTGATCTAGCTCAATCAAACTCTTTTTGCATCATGATACTAGACTAGCTGACTGATCATGACTCTGATCCCGTAGATCGGGTACCTATTACAGTACGATCATCCGATCAGATCATGCTAGTACATCGATCGATACTGCTACTGATCTAGCTCAATCAAACTCTTTTTGCATCATGATACTAGACTAGCTGACTGATCATGACTCTGATCCCGTAGATCGGGTACCTATTACAGTACGATCATCCGATCAGATCATGCTAGTACATCGATCGATACT human genome 3.2 billion bases

Raw genome data

NCBI GenBank Database of genetic sequences gathered from research Publicly available on Web!

Organizing the data

Maps of human genes… Where the genes are… mapping genes & their mutant alleles

Defining a gene… “Defining a gene is problematic because… one gene can code for several protein products, some genes code only for RNA, two genes can overlap, and there are many other complications.” – Elizabeth Pennisi, Science 2003 protein gene RNA gene 1990s -- thought humans had 100,000 genes 2000 -- 40,000 was considered a good estimate 2004 -- 30,000 2006 -- 25,000 is our best estimate gene polypeptide 1 polypeptide 2 polypeptide 3

How does the human genome stack up? Organism Genome Size (bases) Estimated Genes Human (Homo sapiens) 3 billion 30,000 Laboratory mouse (M. musculus) 2.6 billion Mustard weed (A. thaliana) 100 million 25,000 Roundworm (C. elegans) 97 million 19,000 Fruit fly (D. melanogaster) 137 million 13,000 Yeast (S. cerevisiae) 12.1 million 6,000 Bacterium (E. coli) 4.6 million 3,200 Human Immunodeficiency Virus (HIV) 9700 9