carsen group national sales meeting media cybernetics genomics initiative
TRANSCRIPT
Carsen Group National Sales Meeting
Media Cybernetics Genomics Initiative
A Brief History
Perspective
1856- Gregor Mendel begins experimenting with pea plants.
1859- Charles Darwin publishes ‘The Origin of Species’.
1900- DeVries and Correns independently discover and verify Mendel’s principles. Modern era of genetics is born.
1908- Garrod proposes diseases are due to “inborn errors of metabolism” that may result from lack of specific enzymes.
1927- Muller uses x-rays to create artificial mutations in Drosophila.
1950- Chargoff and Franklin lay groundwork for determining basic structure of DNA.
1953- Watson and Crick solve the three dimensional structure of DNA.
1957- Crick proposes the ‘Central Dogma’.
1966- Nirenberg cracks the code that determines how DNA creates proteins.
1988- Human Genome Project begins.
2000- Rough draft of human genome is announced.
What is ‘Genomics’?
Perspective
“Investigations into the structure and function of very largenumbers of genes undertaken in a simultaneous fashion.”1
The field of genomics is undertaking a massive effort to sequencegenomes of several plant and animal species. By understandingthe DNA sequences of these organisms, and the proteins created bythem, we can can better understand the nature of diseases and take steps to prevent those diseases from occurring.
1 Genomics at UC Davis- www.genomics.ucdavis.edu/what
DNA- The Basics
Definitions
DNA- Deoxyribonucleic Acid
Composed of four distinct components; Adenine (A), Cytosine (C),Guanine (G), Thymine (T).
These components are the basic building blocks from which proteinsare created.
Proteins are the structural and regulatory molecules that make eachlife form (grass, people, mouse, bacteria, etc.) unique.
DNA- The Basics
Definitions
A, C, G and T are arranged in sequences which create genes.
Each gene creates a specific protein. Each protein has a function-either structural or regulatory.
The collection of genes within a particular species is referred to asa genome.
Now let’s take a look at how this works...
DNA- The Basics
Definitions
“The Central Dogma”
DNA RNA ProteinTranscription Translation
In the words of Francis Crick:
“Once information has passed into protein, it cannot get out again.”
DNA- The Basics
Definitions
A method for separating fragments of DNA, RNA or protein into discreet groups. The groups create a unique pattern of bandscomposed of fragments of similar molecular size and weight.
Electrophoresis Gels fall into two major classes; Agarose and SDS-PAGE. Agarose gels are typically used to separate DNA fragments,SDS-PAGE are most commonly used to separate proteins.
After separation, it then becomes possible to analyze differentcharacteristics of the molecules being studied. Examples of suchcharacteristics include gene and protein expression.
What is Electrophoresis?
Definitions
Types of Gels and Blots
Applications
Protein Gels- Separate expressed proteins based on their molecular size or charge.
Western Blots- A protein gel derivative.
Southern Blots- Used for determining levels of DNA expression.
Northern Blots- Used for determining levels of RNA expression.
RNA
DNA
Protein
Visualizing Bands- Part I
Applications
Bands are visualized using a variety of markers or ‘tags’.
Proteins- Visible stains such as Coomassie Blue. Also used areradioactive iodine [125I] or sulphur [35S], depending on the application.
DNA/RNA- Fluorescent dyes like Ethidium Bromide (EtBr).Also used are tags such as radioactive phosphorus [32P] or [33P].
Each type of tag has characteristics which make it suitable forthe application.
Visualizing Bands- Part II
Applications
How do you see samples labeled with fluorescence or radioactivity?
• Autoradiography- Exposure to x-ray film.
• Radioisotopic Capture Devices- Direct detection of radioactivity.
• Fluorescence Detection Devices- Scanners, CCD cameras.
How do you quantify the signal?
• By eye- Yes people still analyze autoradiograms by eye!
• Digital Input- Benchtop scanners, and the aforementioned direct detection methods.
Analyzing a Protein Gel
Examples
How much protein is present in a band of interest?
Analyzing a Protein Gel
Examples
What is the molecular weight of the proteins?
What is a Microarray?
Definitions
Microarrays are glass or silicon wafers, or nylon membranes containing a two dimensional matrix of genes. The genes on thewafer are deposited through automated means.
After deposition the genes are probed, typically using a fluorescentlabel, to determine the level of expression generated. These levelscan change depending on the environment to which the genes areexposed, such as a potential drug treatment. By quantifying the change in light intensity for each spot in the microarray, an accuratepicture can be derived for the activity levels for each gene, and how they affect one another during a particular treatment.
Visualizing Spots- Chemistry
Applications
• DNA is spotted onto the previously mentioned array. The DNAexists in the form of cDNA (complementary DNA), which is a mirror-image strand of the section of DNA being studied.
• mRNA is purified from the tissue being studied. The mRNA is thensubjected to a reverse transcriptase procedure which creates a new strand of cDNA. One population of the new strands of cDNA is labeled with the fluorescent tag Cy3, and the other population withCy5.
• The labeled strands are hybridized (‘linked’) to those spotted onto the microarray. Fluorescence excitation yields intensity levels foreach spot in the array. By comparing the amount of signal present inCy3 and Cy5 labeled samples, an estimation of gene expression maybe derived!
Visualizing Spots- Instruments
Applications
• Two major types of instruments are used to collect signal from the microarray. One uses a laser excitation, the other an arc lamp.
• Both types of instruments rely on scanning stages to move thechip under the objective lens. Data are assembled as images andthen the spots assayed for signal.
Modeling Data
Examples
The Genomics Marketplace
Perspective
• Shift toward ‘in silico’ biology- a combination of computersand biology.
• Could be as large as 2 billion USD within next five years.2
• Includes the fields of genomics, proteomics and bioinformatics.
• Who will use this new technology? - Pharmaceutical Companies - Academic Institutions
• What are the applications it will be used for? - Drug Discovery - Crop Protection - ‘Designer Drugs’
2 Scientific American. July 2000.
The Competition
Marketing
Company Website
Affymetrix www.affymetrix.com
Axon Instruments www.axon.com
Alpha Innotech www.alphainnotech.com
Biodiscovery www.biodiscovery.com
Genomic Solutions www.genomicsolutions.com
GSI Lumonics www.gsilumonics.com
Hitachi Genetic Systems www.hitachigeneticsystems.com
Imaging Research www.imaging.brocku.ca
Incyte Genomics www.incyte.com
Research Genetics www.resgen.com
“It has not escaped our notice that the specific pairing we havepostulated immediately suggests a possible copying mechanismfor the genetic material.”
James Watson and Francis Crick- 1953
For the Record
Quotable
Dr. James Watson with a model ofthe DNA double helix.
Dr. Francis Crick dances with hisdaughter at the 1962 Nobel Awardsbanquet.
Resources
For more information
The Beginner’s Guide to Molecular Biology- www.iacr.bbsrc.ac.uk/notebook/courses/guide/
Genomics Education. Celera Science. www.celera.com/celerascience/homepage.cfm?object_name=Genomics%20 Education&parent_menu=Genomics%20Education
Landmarks in the History of Genetics- cogweb.english.ucsb.edu/EP/DNA_history.html
The Biotech Chronicles- www.accessexcellence.org
Genomics: A Global Resource. The Pharmaceutical Research and Manufacturers of America-http://genomics.phrma.org/education/index.html
References
Bibliography
Old R.W. and Primrose S.B. Introduction to Gene Manipulation. 8-13. In Primrose S and Old, R. (Eds.) 1989. Principles of Gene Manipulation. Blackwell Scientific Publications.
Watson J and Crick F. Molecular Structure of Nucleic Acids: A Structure for DeoxyriboseNucleic Acid. Nature (171):737-738.
The Biotech Chronicles- www.accessexcellence.org
Genomics at UC Davis- www.genomics.ucdavis.edu/what.html
Cortese J. Array of Options. Instruments to exploit the DNA microarray explosion. The Scientist14(11): 26.
Gene Genie- Red Herring Magazine (1996). www.herring.com/mag/issue37/gene.html.
Evertsz E, Starink P, Gupta R and Watson D. Technology and Applications of Gene ExpressionMicroarrays. 149-154. In Schena M. (Ed.) 2000. Microarray Biochip Technology. Eaton Publishing, Natick, MA.
Brown, K. The Business of the Human Genome. Scientific American. July 2000. 50-55.