1) Selective breeding
2) Genetic engineering – humans tinker with organisms genes a. Cloning –take haploid cel and replace with a diploid nucleus (comes from organism whos traits you want to duplicate. b. Recombinant DNA – DNA from 2 or more sources. Done by Euk cells during Meiosis. Always from same molecule. 3) Biologists first started doing recombinant DNA from a prok cell and combined it with another prok cell because it was easier. 4) DNA from a euk cell and combine it with DNA from a prok cell Features of bacteria that make them useful subjects for recombinant DNA technicians: 1) Short generation time. Divide fast
2) Easy uptake of DNA from environment (transport from environment) [Fred Griffith transformation did this]. a. Many have Calcium driven DNA pumps. (especially small DNA molecules) b. Electroporation – shock DNA to open pores in plasma membrane so DNA can enter cell c. Have small DNA molecules w/ 0 plasmids 1 ori and carry accessory genes 3) Restriction endonucleases attack DNA molecule from inside. Exonucleases – chop off 1 nucleotide at a time at the ends of the DNA molecule. Not in abundant in prokaryotes Bacterial DNA gets methylated – restriction enzymes cut only unmethylated DNA Bacterial DNA gets bacteria add CH3 to Cytosine or Thymine to protect their own enzymes
Over time they have to choose to be unmethylated or methylated to cut DNA 2 types of restrictiom enzymes used to cut plasmids
Restriction enzymes that open up plasmids at a restriction site
4-8 base pairs long
Make staggered cuts
Plasmids are opened at one restriction site. Foreign Dna treated with same restriction enzymes Naïve bacteria have no plasmids.
Plasmid with wrong foreign DNA fragment
Plasmid with gene of interest
-mix with naïve bacteria get you types of bacteria
Results of recombined plasmid exposed to naïve bacteria- the 4 results: 1: naïve bacteria – will die...