Download Biotechnological Tools and Techniques

Biotechnology
6.1 - Biotechnological Tools &
Techniques
Biotechnology
• the use of living organisms in industrial,
agricultural, medical, and other
technological applications
• there are many biotechnological tools
and techniques used to gain knowledge
of and to manipulate DNA
Restriction Enzymes
• first restriction endonuclease (enzyme)
isolated in 1970
• produced by bacteria
• function as an “immune system” against
invading viruses by cutting up the viral
DNA or RNA
Restriction Enzymes
• restriction enzymes cut double-stranded
DNA at a specific recognition site
• recognition sites are always
palindromic: (same sequence when
read from the 5’ to 3’ direction on either
strand)
Restriction Enzymes
Image from: http://barleyworld.org/css430_09/lecture%208-09/notes8-09.htm
Blunt Ends vs. Sticky Ends
Image from: http://schoolworkhelper.net/2010/07/restriction-endonucleases-or-restriction-enzymes/
DNA Ligase
• DNA ligase can be used to form the
phosphodiester linkages between
fragments of DNA cleaved by restriction
enzymes
• See simple animation:
http://www.dnalc.org/resources/animations/restriction.html
Methylases
Image on next slide: http://www.uic.edu/classes/bios/bios100/lectures/techniques.htm
• enzymes that protect bacterial DNA
from being cleaved by restriction
enzymes
• add methyl groups (-CH3) to one of the
bases in the recognition site, preventing
digestion by restriction enzymes
Plasmids
• circular pieces of non-chromosomal
DNA found in bacteria cells
• artificial plasmids have been engineered
to contain an area with many
recognition sites (and none in other
areas)
Plasmids
• DNA can be inserted into plasmids by
using restriction enzymes and DNA
ligase
• can then introduce these genes into
bacterial cells: transformation
Plasmids
• bacteria are able to express foreign
genes inserted into plasmids, such as
the gene for insulin
• See narrated animation:
http://www.sinauer.com/cooper5e/animation0402.html
Gel Electrophoresis
• a method of separating molecules
based on size
• DNA migrates through the gel towards a
positive electrode
• smaller fragments move faster through
the gel, causing separation by size
• DNA fragments can be seen by using a
stain such as ethidium bromide
Gel Electrophoresis
• Excellent interactive animation of gel
electrophoresis:
http://learn.genetics.utah.edu/content/labs/gel/
• Slideshow with focus on scientists and
history of gel electrophoresis:
http://www.dnalc.org/resources/animations/gelelectroph
oresis.html