Genetic Engineering Techniques (College Board AP Biology)

• Genetic Engineering is a term to describe a number of techniques of manipulating DNA and RNA to achieve certain desired outcomes
• It relies on modifying the characteristics of an organism by manipulating its genetic material
• An important technique within genetic engineering is gel electrophoresis
  • This separates molecules of DNA according to their relative sizes and electrical charge

• Gel electrophoresis is a technique used widely in the analysis of DNA, RNA and proteins
• During electrophoresis, the molecules are separated according to their size/mass and their net (overall) charge
• This separation occurs because:
  • The electrical charge molecules carry:
    • Positively charged molecules will move towards the cathode (negative pole) whereas negatively charged molecules will move towards the anode (positive pole) eg. DNA is negatively charged due to the phosphate groups and thus when placed in an electric field the molecules move towards the anode
  • The different sizes of the molecules:
    • Different sized molecules move through the gel (agarose for DNA and polyacrylamide – PAG for proteins) at different rates
    • The tiny pores in the gel result in smaller molecules moving quickly, whereas larger molecules move slowly
  • The type of gel:
    • Different gels have different sized pores which affect the speed at which the molecules can move through them

DNA separation

• DNA can be collected from almost anywhere on the body, e.g. the root of a hair or saliva from a cup
• After collection DNA must be prepared for gel electrophoresis so that the DNA can be sequenced or analyzed for genetic profiling (fingerprinting)
• To prepare the fragments scientists must first increase (amplify) the number of DNA molecules by the polymerase chain reaction (PCR).
• Then restriction endonucleases (enzymes) are used to cut the DNA into fragments
  • Different restriction enzymes cut the DNA at different base sequences
  • The fragments produced can be characterized by running them along a gel electrophoresis tank

Gel Electrophoresis Diagram

 

The process of electrophoresis