Biotechnology pt1

• Restriction endonucleases


  Enzymes that act like molecular scissors that recognize a specific DNA sequence and cleave the DNA

  
• Reverse transcriptase


  Enzyme that can synthesize DNA from an RNA template
• Taq polymerase


  Enzyme used in PCR to replicate DNA
• Making recombinant DNA


  1. Use restriction endonucleases to cut DNA

  2. Use DNA ligase to join DNA fragments

  
• Vector


  DNA molecule used as a vehicle to carry foreign genetic material into another cell
• Vectors


  • Plasmids
  • Phage
• Designing a plasmid


  1. Include origin of replication

  2. Include restriction site

  3. Include selectable marker

  4. Include differential marker
• Gel electrophoresis


  Technique to separate and purify DNA fragments
• Types of DNA libraries

  • cDNA library
  • Genomic library
• Genomic and cDNA libraries

  Useful in recombinant technology and evolutionary understanding
• Identifying genes of interest

  1. Use hybridization with radioactive probe

  2. Use replica plating or southern blots
• Transforming cells


  Use plasmids with selective and differential markers (Lac Z and GFP)
• Short tandem repeats (STRs)


  Used in DNA analysis and fingerprinting
• DNA fingerprinting using STRs


  Procedure and applications
• CRISPR RNA (crRNA)


  Allows for direct editing of the genome
• DNA sequencing using automated method


  Recognize dideoxy chain terminating nucleotide and its role

  
• Biotechnological procedures


  • Use in forensics
  • Finding evolutionary links
  • Cloning technology
  • Emerging applications
  • Benefits and dangers
• Biotechnology


  Technology that harnesses biological processes to develop products that can help improve our lives
• Ancestral biotechnology started with the invention of agriculture and domestication of plants and animals
• Ancestral biotechnology


  • Domestication of corn
  • Domestication of mustard plants
  • Domestication of chickens
  • Domestication of sheep and cows
  • Food fermentation
• Modern biotechnology combines recent discoveries in molecular biology and genetics with more traditional biotechnologies
• Tools of modern biotechnology

  • Restriction enzymes
  • Electrophoresis
  • Vectors (plasmids, phages)
  • Polymerase chain reaction (PCR)
  • Recombinant DNA

  
• Restriction endonucleases (restriction enzymes)


  Enzymes that recognize a specific DNA sequence and cleave the DNA

  
• Bacteria use restriction enzymes to degrade the DNA of invading bacteriophages

  
• Restriction enzymes


  • They cut the phosphodiester bonds of the phosphate-sugar backbone of DNA
  • Bacteria have methylase enzymes that protect their own DNA from being cut

  
• Types of restriction enzymes

  • Type I and III (recognize sequence but cut randomly)
  • Type II (recognize specific sequence and cut within recognition region)

  
• Palindromic sequence


  DNA sequence that reads the same forward and reverse
• Restriction enzymes


  • EcoR1 (recognizes GAATTC and cuts between G and A)

  
• Sticky ends


  DNA fragments with overhanging ends that can be easily linked
• Blunt ends


  DNA fragments with flat ends after cutting
• Hundreds of different restriction enzymes can be ordered for biotechnology experiments

  
• Restriction fragment length polymorphism (RFLP)


  Technique to distinguish individuals, populations or species based on differences in restriction enzyme sites

  
• Gel electrophoresis


  Technique to separate and purify DNA fragments based on size
• DNA ladder


  DNA fragments of known length used as a reference
• Creating recombinant DNA


  1. Use restriction enzymes to cut DNA

  2. Use DNA ligase to join DNA fragments

  
• DNA ligase


  Enzyme that binds two DNA fragments together
• Recombinant DNA


  • Croco-frog (DNA from crocodile and frog)
• Vectors


  DNA molecules used to carry foreign genetic material into another cell
• Vectors


  • Plasmids
  • Bacteriophages
• Plasmid vectors


  • Have an origin of replication
  • Have a restriction site
  • Can include selectable and differential markers