BIO 205 Microbiology Lecture Outline

Last Modified: February 12, 2002 (J. Crimando).

BIO205 Microbiology
Lecture Outline 6: Microbial Genetics

I. General Genetics

          A. Chromosomes, Genes, DNA, Nucleic Acids
              1. DNA repication
                   a. semiconservative replication
                   b. RNA polymerase, RNA primer, DNA polymerase enzyme, DNA ligase
                   c. general events at the "replication fork"
                   d. leading & lagging strands
                   e. read DNA in 3'to 5' direction; synthesize in 5'to 3' direction.
                   f. energy for DNA synthesis supplied by nucleotides used in synthesis
              2. Major components of DNA and RNA
                   a. nucleosides, nucleotides, nitrogenous bases
                   b. complementarity of bases
                   c. 5' and 3' ends of nucleic acids
               What are the functions of the three types of RNA?
              3. Central dogma of molecular biology
                   a. DNA -> RNA -> Protein
              4. Major factors and events of transcription
                   a. DNA template
                   b. mRNA
                   c. RNA polymerase
                   d. promoter (TATA) & terminator regions of DNA
                   e. exons and introns
              5. Major events of translation
                   a. mRNA template
                   b. ribosome
                   c. ribosome binding site
                   d. tRNAs
                   e. amino acids
                   f. codon on mRNA & anticodon on tRNA
                   g. start (AUG) and stop (UAA, UAG, or UGA) codons
                   h. the concept of the "Genetic Code"
           In a eukaryotic cell, where do transcription and translation occur?
          B. Regulation of Metabolism
              1. Regulation of enzyme activity (already covered previously)
              2. Genetic regulation of enzyme production
                   a. Induction (common in catabolism regulation)
                         i. Example: lac Operon
                               structural genes
                               regulatory sites (promoter & operator regions)
                               inducer molecule (lactose)
                               repressor protein
                               regulator (i) gene
                               * lactose inactivates the repressor, thus inducing the lac genes
                   b. Repression (common in anabolism regulation)
                          i. Example: trp Operon
                               tryptophan activates a repressor protein, thus allowing repression of the trp genes
           Why is induction (rather than repression) often used to regulate catabolic enzymes?
           Why is repression (rather than induction) often used to regulate anabolic enzymes?
          C. Mutations
              1. Mutagen
              2. Types of mutations
                    a. point mutations (substitutions/replacements)
                           THE ONE BIG FLY HAD ONE RED EYE
                          THE ONE BIG FLY HAS ONE RED EYE
                          THE ONE BIG FLP HAD ONE RED EYE
                    b. deletions/insertions (frameshift mutations)
                          THE ONE BIG FLY HAD ONE RED EYE
                          THE ONE BGF LYH ADO NER EDE YE
                          THE OEE EEB IGF LYH ADO NER EDE YE
                Which is more detrimental, point or frameshift mutations? Why?

II. Biotechnology

A. Basic Terminology (Know definitions):

   1. Restriction enzyme
         Example: EcoR1
   2. Restriction site
         Example: ...G^vAATTC...
   3. Vector
         Example: E.coli plasmid
   4. Clone
   5. "Sticky ends"
   6. Genetic Recombination
   7. DNA Fingerprint
   8. Restriction Fragment Length Polymorphism (RFLP)
   9. Southern Blotting
   10. Polymerase Chain Reaction (PCR)
   11. Primer

B. Gene Cloning
Describe General Procedures Including:

   1. Digestion of DNA using restriction enzymes,
   2. Insertion into vector,
   3. Uptake and expression of recombinant gene in E. coli
   4. Identify some examples of using gene cloning

C. DNA Fingerprinting
Describe General Procedure Including:

   1. Digestion of DNA using restriction enzymes
   2. Loading/running into gel for fragment separation
   3. Identification of different size fragments in gel (RFLP analysis)
   4. Comparison of "banding" patterns - "DNA Fingerprint"
   5. Southern blotting transfer of separated fragments onto nitrocellulose paper
   6. Use of radiolabelled probe (short single stranded sequence complementary to gene of interest)
       to hybridize to and identify which band contains a selected gene or DNA sequence

D. Polymerase Chain Reaction (PCR)
Describe general procedure including:

1. Heat denaturation of DNA
2. Hybridization of short primers to DNA
3. Replication of target strand using heat-resistant DNA polymerase (Taq)
4. Repeated cycling to amplify target DNA

This is only a general outline.
There may be material that has been discussed in lecture that is not included in this outline
and there may be material on this outline that has not been discussed in lecture.
Any material discussed in lecture or listed in this outline is "fair game" for the test.