1. 6.1 - Biotechnological Tools & Techniques
Biotechnology
6.1 - Biotechnological Tools & Techniques

2. 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

3. 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

4. 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)

5. Restriction Enzymes Image from: http://barleyworld

6. Blunt Ends vs. Sticky Ends Image from: http://schoolworkhelper

7. DNA Ligase
DNA ligase can be used to form the phosphodiester linkages between fragments of DNA cleaved by restriction enzymes
See simple animation:

8. Methylases Image on next slide: http://www. uic
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

10. 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)

11. Plasmids
DNA can be inserted into plasmids by using restriction enzymes and DNA ligase
can then introduce these genes into bacterial cells: transformation

12. Plasmids
bacteria are able to express foreign genes inserted into plasmids, such as the gene for insulin
See narrated animation:

13. 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

14. Gel Electrophoresis
Excellent interactive animation of gel electrophoresis:
Slideshow with focus on scientists and history of gel electrophoresis: