Lecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityBacillus anthracisLast Lecture:1. Introduction2. History3. Koch’s PostulatesToday’s Lecture:Today s Lecture:1. Prokaryote vs. Eukaryote2. Classifying prokaryotes3Ph l ti3.PhylogeneticsLecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityI. Basic Cell structure: (Fig. 1.5)A. All cells have B Many cells (plants prokaryotes fungi) have:B. Many cells (plants, prokaryotes, fungi) have: C. Eukaryotic cells 1. Major feature that is not in prokaryotic cells are: • • • 2. Generally larger and more complex than prokaryotes (Fig. 23.1) 3. DNA is contained in a __________________ compartment called the _______________ D. Prokaryotic cells (Fig. 4.43) 1. Two domains: ____________ and ________________ 2. Generally ____________ and _____________than eukaryotes 3. DNA is free in the cytoplasm.Lecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityII. Characteristics of the Domains of LifeCharacteristics of the primary domains (Table 1.2, 1.3, 1.4) Property BacteriaArchaea Eukaryote p yyNuclear membrane Peptidoglycan cell walls Membrane Lipids: glycerol Lipids: glycerol-hydrocarbon linkage Contain plastids Mitochondria Chloroplasts Transcription mRNA processing (capping, polyA t il) tail) mRNA splicing RNA Polymerase Genes in operons Transcription Factors Required Ribosome sizeRibosome size Metabolism: Nitrogen Fixation Chemolithotrop hy Growth > 80˚C Should be ester linkage NOT etherLecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityIII. Artificial vs. Phylogenetic Classification A. Artificial classification is based on: ________________________. E.g. g B. Phylogenetic classification is based on: ______________________.1. Requires: 2 Preferred by bacteriologists2. Preferred by bacteriologistsC. Two prokaryotic domains: IV. Nomenclature follows the binomial system of names. A. Domain, Phylum, Class, Order, Family, Genus, Species (Table 17.1) B. Example: Genus, Species: Escherichia coli must be Latin endings. 1. Genus is always capitalized and the species is lower case 2 Al it li i d li2. Always italicize or underline.3. Name usually has some significance. C. How do identify a new isolate and classify it to the species level? 1. There are international guidelines: e.g. DNA-DNA hybridization IV. Phylogenetic Trees A. Basically two ways to create a phylogenic tree: 1. using: 2. using: B. The molecular-based system 1. Phylogenetic Tree shown in Fig 1.6 a) )b) The tree is derived from c) Pioneered by ________________________(Box 17.4) 2. This organization suggests that most of the diversity of life is in the domains based on ribosomal RNA differencesthe ____________domains based on ribosomal RNA differences.a) Humans to corn: _________base changes/site. b) Between bacteria and archaea: ___________base changes/site. 3. Archaea are __________ closely related to Eukarya than Bacteria.Bacteria. 4. Eukaryotic microbesLecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityVI. The 16S or 18S rRNA gene is commonly used for phylogenetic classification of prokaryotes and eukaryotes:classification of prokaryotes and eukaryotes:A. General features of a molecular marker? 1. 2. The function is: 3. There are ____________ and _______________regions 4. The gene tolerates: B. The gene encoding 16S ribosomal RNA (18S in eukaryotes) 1. The 16S rRNA is part of the ribosome a) Occurs in _______________________ b)b) c) Gene size: d) composed of many domains that can change independently of each other (Figure 17.1—SEQUENCED BY HARRY NOLLER) e) f) Large databases of sequences (Ribosome Database Project) VII. Methods for making a phylogenetic treeA. Phylogenetic trees: Figure 17.4 and 17.5, PCR and sequencing discussed in Boxs 16.1 and 16.3 1. Structure: nodes (internal vs. external), branches, and branch lengths (17.4) 2 T i t t d d t d (17 3)2. Two main types: rooted and unrooted (17.3)Lecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityB. Steps to make a 16S rRNA gene Phylogenetic Tree 11. 2. Use Polymerase Chain Reaction (PCR) to: What goes into a PCR: 1) 2) 3) 4) )5) Put tubes in a thermocycler machinge. What does this do? Denature:Denature: Anneal: Extension: 3. Analyze by agarose gel electrophoresis 4. Sequence the 16S rRNA gene and compare the sequence to a database of sequences to see what prokaryote you’ve got by ALIGNING yours to others. 5. Turn the alignments into a: 5 u t e a g e ts to a__________________________Let’s go over Fig. 17.6Lecture 2 Bio119Prokaryotes vs. Eukaryotes and Diversity!"C l l t di t t i!"Calculate distance matrix!"Start an intermediate tree with most closely related strains.!"Calculate second distance matrix.!"Add other intermediate tree of closely related strains!"Calculate distance between two intermediate trees!"Calculate distance between two intermediate trees!"Then the average distance and draw the branchesd(ab)(cd)Lecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityFig. 1.5, 23.1Lecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityFig. 4.43, 1.6Lecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityFig. 17.1Lecture 2 Bio119Prokaryotes vs. Eukaryotes and DiversityBox