Genetic Engineering Step 1 Study Guide

  • What characteristics make plasmids easy to genetically modify?

    • Plasmids have short sequences of DNA
    • Plasmids are easy to cut and paste back together
    • Plasmids copy independently

  • Explain the process and all reagents used to extract DNA.

  

  1. Lysozyme is added degrade peptidoglycan in cell walls
    • We don’t use lysozyme because we have animal cells
  2. Detergents are added to lyse (explode) the cell
    • Detergents include sarkosyl or sodium dodecyl sulfate (SDS)
    • Proteases and salts are added to clump debris
  3. The mix is then centrifuged to create a precipitate and a supernate

      1. The precipitate is at the bottom and is debris 2. The supernate is at the top and has the DNA 3. DNA can be extracted with alcohol

         

  • Explain how PCR and Gel Electrophoresis are used in genetic engineering.
    • PCR- Stands for Polymerase Chain Reaction, used to copy DNA many times
    • PCR uses primers which target specific sequences in DNA
    • DNA strands are split apart at 95°
    • Then primers bind to the split strands at 50°
    • The polymerase enzyme makes a complementary strand using free nucleotides at 72°

       

  • Gel Electrophoresis- Separates DNA fragments based on size
    • There is a gel made of agarose that has wells (holes) in it
    • DNA is placed into the wells
    • An electric charge is run through the gel
    • DNA is negatively charged and it will move to the positively charged side
    • Smaller fragments will move faster so they will go further

Vocabulary:

genetic engineering- manipulation of genetic information

multiple cloning sites- series of unique restriction enzyme recognition sites

vectors- Vessel to carry genetic information with (often a plasmid)

lysosome- degrades peptidoglycan in cell walls

precipitant- molecular debris, found at bottom of test tube

supernatant- suspends DNA, found at top of test tube

lysed- to explode cells

detergent- Solutions to lyse cells with

protease- Degrades proteins

salts- clumps cellular debris

probes- single-strands of DNA or RNA that bind to a complementary sequence that have a biological marker attached. Shows location of DNA sequences.

hybridization- Process where probe binds to complementary sequence

restriction enzymes- cuts DNA at specific sites

gel electrophoresis- separates DNA fragments based on size

DNA polymerase- Synthesizes new DNA strands in the PCR process

thermal cycler- Used in PCR to take DNA through temperatures best for replication (95°, 50°, 72°)

polymerase chain reaction- used to replicate specific sequence of DNA many times

  • We do not replicate all of the DNA

southern blotting- transfers DNA from gel onto membrane with probes. Uses photographic film to identify specific sequences of DNA

exons- coding sequences that leave the nucleus

introns- non-coding sequences of DNA that cannot leave the nucleus