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Molecular Biology Topics for Second Part of Molecular Biology Course: Structural Organization of Eukaryotic Chromosomes Spring 2009 BI 425 UOG.

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Presentation on theme: "Molecular Biology Topics for Second Part of Molecular Biology Course: Structural Organization of Eukaryotic Chromosomes Spring 2009 BI 425 UOG."— Presentation transcript:

1 Molecular Biology Topics for Second Part of Molecular Biology Course: Structural Organization of Eukaryotic Chromosomes Spring 2009 BI 425 UOG

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21 NonTranscribed genes are less susceptible to Dnase 1 Digestion

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25 Fluorescent Labeled Probes hybridized to Interphase Chromosomes

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32 Prokaryotic Gene Regulation

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35 Deletion Analysis: Identify Upstream Transcription Control Sequences

36 Determination of Consensus TATA Box Sequence

37 Scanning Mutations Identify Transcription Control Elements

38 General Pattern of Control Elements that Regulate Gene Expression in Yeast and other Eukaryotes.

39 LIFE CYCLE OF HIV-1 RNA DNA DNA Provirus Latency Viral Transcription Viral RNA RNA Processing mRNA Viral Proteins Viral genomic RNA Virion Assembly Maturation & Release of Virus HIV Virus 3’ LTR pol env nef if vpr vpu rev Shubir Ghosh, UCSF

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41 Overview of Eukaryotic Transcriptional Control

42 Column Chromatography Separation of the three RNA Polymerases In Eukaryotes.

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45 Transcription Assay: Transcription Start Site

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47 EMSA

48 Dnase 1 Footprinting

49 Dnase1 Footprint Assay in Transcription Factor Purification

50 Northern Blot & S1 Nuclease Assay: Identify Enhancers of Transcription S1 Nuclease Assay Northern Blot Homework!

51 In vitro Transcription Assay: Identification of Transcription Factors

52 In vivo Transfection Assay: Identify & Evaluate Transcription Factors

53 Eukaryotic Transcriptional Activators: Features & Experimental Analysis

54 Eukaryotic Transcriptional Activators: Features & Experimental Analysis of GAL4

55 Eukaryotic Transcriptional Activators: Features & Experimental Analysis of GAL4

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58 Interaction of bacteriophage 434 Repressor with DNA

59 Interaction of Zinc Finger proteins with DNA GR

60 Interaction of Lecuine Zipper & basic HLH proteins with DNA Yeast Gcn4 Transcription Activator

61 Cooperative binding of two unrelated transcription factors to neighboring sites in a composite control element

62 Combinatorial Possibilities due to formation of Heterodimeric Transcription Factors

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65 Transcriptional Repression : Yeast mating Locus

66 Antibody and DNA probes colocalize RepressorProtein, Sir3, with telomeric heterochromatin in Yeast nuclei Telomeres labeled by Hybridization to Telomere Specific DNA probe (Yellow) Telomere specific DNA Hybridization Probe Fluorescent labeled Antibody specific For Sir3 protein

67 Schematic Model of Silencing Mechanism at Yeast Telomeres Dependence of Silencing on Hypoacetylation of the Histone Tails

68 Handout on Histone Acetylation/Deacetylation! Model of Transcriptional Activation Mechanism See HACs & HDACs

69 Chromatin Immunoprecipitation Assay: Evaluate State of Histone Acetylation on Nucleosome

70 Transcriptional Activation by Gcn4 in Saccharomyces cerevisae Gcn4 UAS Gcn5 + Multisubunit Complex Gcn4 Activated Gene in S.c Components of Gcn4 activated gene in yeast:

71 Mechanism of Transcriptional Activation by Gcn4

72 Mechanism of Transcriptional Repression by Ume6 in Yeast Components of Repression in yeast by Ume6: Ume6 Repressed gene in S.c Ume6 Multisubunit complex including: Sin3 Rpd3

73 Mechanism of Transcriptional Repression

74 Medistor Complex in Euksryotic Transcription

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