6: techniques

• rocombinant dna is DNA put together from different sources
• plasmid vectors are used to clone genes
• host cells are transformed with plasmids
• making bacteria competent allows it to take up foreign DNA
• plasmids are small circular DNA molecules that are found in the cytoplasm of bacteria
• the transformant is the cell that has taken up the plasmid
• a cloning vector is a piece of DNA that is stable enough to take in foreign DNA
• an idea cloning vector:

  • needs an origin of replication
  • needs selectable markers
  • needs recognition sites for restriction enzymes
• selectable markers are genes that have a visual ot growth selective marker
• isogenic bacterial cultures: bacteria that have the same genetic makeup
• restriction enzymes: enzymes that cut DNA at specific sequences of bases
• when the DNA is cut, it is digested
• T4 DNA ligase is a DNA polymerase that joins DNA fragments together by adding a phosphate group to the 3' end of the new strand
• restriction sites: sequences of DNA that are recognized by restriction enzymes and cut when they are encountered
• Kary Mullis developed PCR
• the first step of the PCR is to denature the DNA by heating it to 95 degrees for a few minutes
• the second step of the PCR is to attach primers to the single stranded DNA