© life_edu lecture 4 some techniques in biotechnology issues in biotechnology: the way we work with...
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© life_edu
Lecture 4
Some Techniques in Biotechnology
Issues in Biotechnology:The Way We Work With Life
Dr. Albert P. Kausch
life edu.us
The Mechanics of DNA
Issues in Biotechnology:Biotechnology, Our Society and Our Future
OnCampus LiveOnCampus LiveBCH 190, MIC 190, AFS 190, NRS 190, PLS 190BCH 190, MIC 190, AFS 190, NRS 190, PLS 190
OnLine BCH 190OnLine BCH 190
A Sweeping General Survey on Life and BiotechnologyA Public Access College Course
The University of Rhode Island
Kimberly Nelson
Issues in Biotechnology:The Way We Work With Life
Dr. Albert P. Kausch
life edu.us
© life_edu
A Sweeping General Survey on Life and Biotechnology
The University of Rhode Island
Issues in Biotechnology:The Way We Work With Life
Dr. Albert P. Kausch
life edu.us
BCH 190BCH 190Section I.
The Mechanics of Life and General Biotechnology
© life_edu
3. Atoms, Cells and the Flow of Life
4. Some Techniques in Biotechnology
A Sweeping General Survey on Life and BiotechnologyA Public Access College Course
The University of Rhode Island
Issues in Biotechnology:The Way We Work With Life
Dr. Albert P. Kausch
life edu.us
The Mechanics of DNA
© life_edu
Lecture 4
Some Techniques in Biotechnology
Issues in Biotechnology:The Way We Work With Life
Dr. Albert P. Kausch
life edu.us
The Mechanics of DNA
Tools of the TradeThe eppendorf tubeand the pipetmanare the standard stock and trade in the dailywork of a molecularbiologist
Tools of the TradeThe eppendorf tubeand the pipetmanare the standard stock and trade in the dailywork of a molecularbiologist
“On the body of the traditional P-Series pipet it says, in relief, “Gilson.” Warren Gilson, who earned his MD in 1940 at the Univ. of Wisconsin, invented and patented the mechanical basis for the popular adjustable pipet (US Patent No. 3,827,305, 1974), Nearly 40 yrs. After the patent, the Pipetman continues to be manufactured in France in a factory started by Gilson’s colleague, Eugene Marteau D’Autry. Shortly before Gilson’s patent issued Gilson sold the marketing and sales rights to Ken Rainin President of Rainin Instrument, because, Gilson says, ‘He was a good salesman.’”
“Eppendorf tubesand PipetmanFor the Gold Rush”
Innovative technologies become biotech products
Separation Techniques: The need to separate the components of Life
Precipitation/DissolutionFiltersCentrifugationAffinityBlotsMagneticsElectrophoresisEtc.
The ultracentrifuge is a centrifuge optimized for spinning a rotor at very high speeds, capable of generating acceleration as high as 2,000,000 G (approx 19 600 km/s²). Ultracentrifuges find important uses in molecular biology, biochemistry and polymer science, including separation of cellular structures and molecules.
Electro refers to the energy of
electricity. Phoresis, from the
Greek verb phoros, means
“to carry across.” Thus, gel
electrophoresis refers to the
technique in which molecules
are forced across a span of gel,
motivated by an electrical
current.
Gel Electrophoresis:the separation of molecules,DNA, RNA and proteinsby charge and size
What is a Gel?What is a Gel?
Agarose is a long chain of sugar molecules,
a polymer, derived from algae
used in electrophoresis to separate molecules
Two types of gel:• Agarose (horizontal type)
• Polyacrylamide (vertical type)
How are Gels Loaded and Run?How are Gels Loaded and Run?
Applications of Gel Electrophoresis
• DNA Fingerprinting
• DNA Recombinant Technology
• Forensics
• The Human Genome Project
DNA carries a net negative charge; it is negatively charged because the phosphates (red circles) that form the sugar-phosphate backbone of a DNA molecule have a negative charge.
The gel matrix acts as a sieve for DNA molecules. Large molecules have difficulty getting through the holes in the matrix. Small molecules move easily through the holes. Because of this, large fragments will lag behind small fragments as DNA migrates through the gel.
As the separation process continues, the separation between the larger and smaller fragments increases.
• Molecular weight markers are often electrophoresed with DNA.• Molecular weight markers are usually a mixture of DNAs with known molecular weights.• Molecular weight markers are used to estimate the sizes of DNA fragments in a DNA sample.
Issues in Biotechnology
Gel electrophoresis is an important tool in molecular biology and biotechnology. Electro refers to the energy of electricity. Phoresis, from the Greek verb phoros, means “to carry across.” Thus, gel electrophoresis refers to the technique in which molecules are forced across a span of gel, motivated by an electrical current. Gel electrophoresis allows for:
(A) the separation of biological molecules, including DNA, RNA and
proteins by their charge and size (B) all of the answers are correct (C) the identification of DNA markers now commonly used in forensics to implicate or exonerate persons accused of various crimes (D) the rapid visualization of the products of PCR (E) the acceleration of DNA into cells for genetic engineering purposes
The Techniques of The Techniques of Molecular BiotechnologyMolecular Biotechnology
Technology has created new Fields
DNA detection GenomicsDNA synthesis Bioinformatics
DNA sequencing Pharmacogenomics
DNA cloning Transgenics
Expression cassette Computationalconstruction Biology
RNA detection Population Genetics
Protein detection Proteomics
The Techniques of The Techniques of Molecular BiotechnologyMolecular Biotechnology
Technology has created new Fields
DNA detection GenomicsDNA synthesis Bioinformatics
DNA sequencing Pharmacogenomics
DNA cloning Transgenics
Expression cassette Computationalconstruction Biology
RNA detection Population Genetics
Protein detection Proteomics
DNA does not replicate spontaneously, but is facilitated by a group of proteins Interestingly, each of these proteins is
coded for in DNA they also replicate
Proteins Are Used to Copy DNA
Enzymes were discovered that cut DNA at specific sequences
And subsequently, enzymes were discovered that paste DNA together
The ability to cut and paste DNA allowed gene cloning
Plasmids are circular pieces of DNA found in some bacteria
Many copies per cell
Antibiotic resistance gene
Plasmids can be cut and pasted back together
Foreign genes can be inserted
How is a gene cloned?Foreign DNA (gene)is inserted into a plasmid that has a gene for antibiotic resistance
The plasmid is introduced into a bacterial cell and grown on the antibiotic
Only bacteria with the plasmid grow…the inserted gene is copied many times
Promoter Coding Sequence Terminator
Gene Construction
Cell specificityDevelopmental specificityStart transcription
Protein coding sequenceStop transcriptionMessage stability
Gene constructs can be moved into plants and the gene is expresseddriven by the promoter sequence
It is now possible to clone any gene from any
organism and move it into any other organism
Gene transfer from one organism to another is not new
Image of two species of bacteria transferring viralphage particles
Bacteria transfer genesto other bacteria and plants
Now in nature thereis another organismcapable of transferring DNA: we call that organisma human being
Tools and Techniques Tools and Techniques used in Biotechnologyused in Biotechnology
No Walls
The Clear bead at the center changes everythingThere are no edges to my being now
I have heard it said that there is a windowThat opens from one mind to another
But where there are no wallsThere is no need for a window, or fitting a latch.
Rumi 1279 AD
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Thank You to The University of Rhode Island Thank You to The University of Rhode Island and all of the students of Issues in and all of the students of Issues in
Biotechnology over the yearsBiotechnology over the years
Dr. Albert KauschDr. Albert Kausch
Dr. Albert Kausch Dr. Albert Kausch and Kimberly Nelsonand Kimberly Nelson
Thaddeus WeaverThaddeus Weaver