Very Basic Biotechnology Supplemental instruction Designed by Pyeongsug Kim ©2010 [email protected]@att.net Fall 2010 For Dr. Wright’s Bio 7/27 Class.

VeryVery Basic Biotechnology Basic BiotechnologySupplemental instruction

Designed by Pyeongsug Kim ©2010 [email protected]

Fall 2010For Dr. Wright’s Bio 7/27 Class

Biotechnology review Define: recombinant DNA technology; genetic engineering; gene cloningHow do restriction enzymes work? How are they used for cloning? What is an expression vector?How does the polymerase chain reaction (PCR) work?What is a probe? What are some possible uses for probes?

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BiotechnologyBiotechnology-The use of microbiological and biochemical techniques to solve practical problems and produce more useful(desirable) products.

Picture from http://www.plantsci.cam.ac.uk/Haseloff/teaching/PlantBiotech/index.html

Genetic engineeringGenetic engineering

-Alternating an organism’s genetic engineering using in vitro techniques.

A procedure performed in vitro (Latin: within the glass) is performed not in a living organism but in a controlled environment, such as in a test tube or Petri dish.

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From http://en.wikipedia.org/wiki/In_vitro

Human embryos photographed developing in vitro

Picture from http://employees.csbsju.edu/hjakubowski/classes/ch331/dna/plasmid.gif

Using microbes(usually bacteria)Using microbes(usually bacteria)-Protein production-DNA production-Gene regulation research-”DNA library”-Why microbes? Grow fast (even single-celled eukaryotes like yeast) Don’t take up much space Economical source of genetically-engineered material Have the same DNA as all living organisms.(DNA or RNA)

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Recombinant DNA technologyRecombinant DNA technology: DNA molecule created by joining DNA fragments from two different sources (two different organisms).

Picture from http://citnews.unl.edu/hscroptechnology/html/printLesson.shtml?lessonID=990631835 Designed by Pyeongsug Kim, ©2010 www.science-i.com

________________-naturally occurring bacterial enzyme -recognizes and cut a specific nucleotide sequence of DNA.-used to cut DNA at specific points during production of recombinant DNA.-ECoRI ~ restriction enzyme from E.Coli(RY13)

-can be joined to fragments to other source.

For recombinant DNA, first, the DNA needs to be cut!And glue it!

Restriction enzymes

Picture from http://employees.csbsju.edu/hjakubowski/classes/ch331/dna/plasmid.gif Designed by Pyeongsug Kim, ©2010 www.science-i.com

Originally the bacterial enzyme that stops viral reproduction by cleaving viral DNA.

Expression vectors

Picture from http://www.artes-biotechnology.com/service/hansenula.jsp

_______________-Generally a plasmid-used to introduce a specific gene into a target cell.-facilitate transcription and translation of cloned DNA.

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Picture from http://www.artes-biotechnology.com/service/hansenula.jsp

Gene cloningGene cloning -Fragments can be “cloned” into “expression vectors” to be inserted into microbes -the cloned cDNA is incorporated into an expression vector or plasmid and transferred into bacterial or yeast cells.The cells multiply (cloning)

The interesting DNA needs to be cloned.

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Cloning stepCloning step -Isolate DNA. -Use a restriction enzyme to generate fragment. -Generate a recombinant molecule. -Introduce recombinant molecule. -Introduce recombinant into new host.

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To express the protein, the plasmid must be placed inside a cell, either prokaryote or eukaryote.

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Is there my interesting DNA?___________-Single-stranded piece of DNA tagged with a detectable marker -used to detect its its complementcomplement. -Analysis of DNA within the cell (germline)

DNA Probes

<Germline> A lineage of cells from which gametes are derived. Also known as germ track

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______________________-Technique used to exponentially amplify specific regions of a DNA molecule.

(PCR)Polymerase chain reaction

Picture from http://blogs.newamericamedia.org/photo-of-the-day/103/pcr http://nobelprize.org/nobel_prizes/chemistry/laureates/1993/illpres/pcr.htmlDesigned by Pyeongsug Kim, ©2010 www.science-i.com

______________________-Can be used to detect the presence of DNA sequence.-Diagnostics, forensic analysis

Polymerase chain reaction (PCR)

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