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BEGR 424 Molecular Biology William Terzaghi Spring, 2014

BEGR 424 Molecular Biology William Terzaghi Spring, 2014. BEGR424- Resource and Policy Information Instructor: Dr. William Terzaghi Office: SLC 363 / CSC228 Office hours: MWF 12:00-1:00, TR 1-2 or by appointment Phone: (570) 408-4762 Email: terzaghi@wilkes.edu.

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BEGR 424 Molecular Biology William Terzaghi Spring, 2014

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  1. BEGR 424 Molecular Biology William Terzaghi Spring, 2014

  2. BEGR424- Resource and Policy Information Instructor: Dr. William Terzaghi Office: SLC 363/CSC228 Office hours: MWF 12:00-1:00, TR 1-2 or by appointment Phone: (570) 408-4762 Email: terzaghi@wilkes.edu

  3. BEGR424- Resource and Policy Information Instructor: Dr. William Terzaghi Office: SLC 363/CSC228 Office hours: MWF 12:00-1:00, TR 1-2 or by appointment Phone: (570) 408-4762 Email: terzaghi@wilkes.edu Course webpage: http://staffweb.wilkes.edu/william.terzaghi/BEGR424.html

  4. General considerations What do you hope to learn?

  5. General considerations • What do you hope to learn? • Graduate courses • learning about current literature

  6. General considerations • What do you hope to learn? • Graduate courses • learning about current literature • Learning how to give presentations

  7. General considerations • What do you hope to learn? • Graduate courses • learning about current literature • Learning current techniques

  8. General considerations • What do you hope to learn? • Graduate courses • learning about current literature • Learning current techniques • Using them!

  9. Plan A • Provide a genuine experience in using cell and molecular biology to learn about a fundamental problem in biology. • Rather than following a set series of lectures, study a problem and see where it leads us. • Lectures & presentations will relate to current status • Some class time will be spent in lab & vice-versa • we may need to come in at other times as well

  10. Plan A Pick a problem Design some experiments

  11. Plan A Pick a problem Design some experiments See where they lead us

  12. Plan A Pick a problem Design some experiments See where they lead us Grading? Combination of papers and presentations

  13. Plan A • Grading? • Combination of papers and presentations • First presentation:10 points • Research presentation: 10 points • Final presentation: 15 points • Assignments: 5 points each • Poster: 10 points • Intermediate report 10 points • Final report: 30 points

  14. Plan A Topics? Cloning and analyzing oxalate decarboxylase and/or oxalate oxidase to see if they dissolve kidney stones in collaboration with Dr. VanWert Making vectors for Dr. Harms Cloning & sequencing antisense RNA Studying ncRNA Something else?

  15. Plan A Assignments? identify a gene and design primers presentation on new sequencing tech designing a protocol to verify your clone presentations on gene regulation presentation on applying mol bio Other work draft of report on cloning & sequencing poster for symposium final gene report draft of formal report formal report

  16. Plan B • Standard lecture course, except: • Last lectures will be chosen by you -> electives

  17. Plan B • Standard lecture course, except: • Last lectures will be chosen by you -> electives • Last 4 labs will be an independent research project

  18. Plan B • Standard lecture course, except: • Last lectures will be chosen by you -> electives • Last 4 labs will be an independent research project • 20% of grade will be “elective” • Paper • Talk • Research proposal • Poster • Exam

  19. Plan B schedule- Spring 2014 • Date TOPIC • JAN 13 General Introduction • 15 Genome organization • 17 Cloning & libraries: why and how • 20 DNA fingerprinting • 22 DNA sequencing • 24 Genome projects • 27 Studying proteins • 29 Meiosis & recombination • 31 Recombination • FEB 3 Cell cycle • 5 Mitosis • 7 Exam 1 • 10 DNA replication • 12 Transcription 1 • 14 Transcription 2 • 17 Transcription 3

  20. 19 mRNA processing 21 Post-transcriptional regulation 24 Protein degradation 26 Epigenetics 28 Small RNA MAR 3 Spring Recess 5 Spring Recess 7Spring Recess 10 RNomics 12 Proteomics 14 Exam 2 17 Protein synthesis 1 19Protein synthesis 2 21Membrane structure/Protein targeting 1 24 Protein targeting 2 26 Organelle genomes 28Mitochondrial genomes and RNA editing 31Nuclear:cytoplasmic genome interactions

  21. APR 2Elective 4Elective 7 Elective 9 Elective 11 Elective 14 Elective 16 Elective 18Easter 21Easter 23 Elective 25 Elective 28Exam 3 30 Elective Last Class! ??? Final examination

  22. Lab Schedule • Date TOPIC • Jan 14 DNA extraction and analysis • 21 BLAST, etc, primer design • 28 PCR • Feb 4 RNA extraction and analysis • 11 RT-PCR • 18 qRT-PCR • 25 cloning PCR fragments • Mar 4Spring Recess • 11 DNA sequencing • 18 Induced gene expression • 25 Northern analysis • Apr1 Independent project • 8 Independent project • 15 Independent project • 22 Independent project

  23. Genome Projects Studying structure & function of genomes

  24. Genome Projects • Studying structure & function of genomes • Sequence first

  25. Genome Projects • Studying structure & function of genomes • Sequence first • Then location and function of every part

  26. Genome Projects • How much DNA is there? • SV40 has 5000 base pairs • E. coli has 5 x 106 • Yeast has 2 x 107 • Arabidopsishas 108 • Ricehas 5 x 108 • Humans have 3 x 109 • Soybeans have 3 x 109 • Toads have 3 x 109 • Salamanders have 8 x 1010 • Lilies have 1011

  27. Genome Projects • C-value paradox: DNA content/haploid genome varies widely

  28. Genome Projects • C-value paradox: DNA content/haploid genome varies widely • Some phyla show little variation: • birds all have ~109bp

  29. Genome Projects • C-value paradox: DNA content/haploid genome varies widely • Some phyla show little variation: • birds all have ~109bp • mammals all have ~ 3 x 109 bp

  30. Genome Projects • C-value paradox: DNA content/haploid genome varies widely • Some phyla show little variation: • birds all have ~109bp • mammals all have ~ 3 x 109 bp • Other phyla are all over: • insects and amphibians vary 100 x

  31. Genome Projects • C-value paradox: DNA content/haploid genome varies widely • Some phyla show little variation: • birds all have ~109bp • mammals all have ~ 3 x 109 bp • Other phyla are all over: • insects and amphibians vary 100 x • flowering plants vary 1000x

  32. C-value paradox • One cause = variations in chromosome numbers and ploidy • 2C chromosome numbers vary widely • Haplopappus has 2

  33. C-value paradox • One cause = variations in chromosome numbers and ploidy • 2C chromosome numbers vary widely • Haplopappus has 2 • Arabidopsis has 10

  34. C-value paradox • One cause = variations in chromosome numbers and ploidy • 2C chromosome numbers vary widely • Haplopappus has 2 • Arabidopsis has 10 • Rice has 24 • Humans have 46 • Tobacco (hexaploid) has 72 • Kiwifruit (octaploid) have 196

  35. C-value paradox Chromosome numbers vary So does chromosome size!

  36. C-value paradox Chromosome numbers vary So does chromosome size! Reason = variation in amounts of repetitive DNA

  37. C-value paradox Chromosome numbers vary So does chromosome size! Reason = variation in amounts of repetitive DNA first demonstrated using Cot curves

  38. Cot curves • denature (melt) DNA by heating

  39. Cot curves • denature (melt) DNA by heating • dissociates into two single strands

  40. Cot curves • denature (melt) DNA by heating • Cool DNA

  41. Cot curves • denature (melt) DNA by heating • Cool DNA:complementary strands find each other &anneal

  42. Cot curves • denature (melt) DNA by heating • Cool DNA:complementary strands find each other &anneal • hybridize

  43. Cot curves • denature (melt) DNA by heating • Cool DNA:complementary strands find each other &anneal • Hybridize: don't have to be the same strands

  44. Cot curves • denature (melt) DNA by heating • Cool DNA:complementary strands find each other &anneal • Hybridize: don't have to be the same strands • Rate depends on [complementary strands]

  45. Cot curves • 1) denature DNA • 2) cool DNA • 3) at intervals measure • [single-stranded DNA]

  46. Cot curves viruses & bacteria show simple curves Cot is inversely proportional to genome size

  47. Cot curves eucaryotes show 3 step curves Step 1 renatures rapidly: “highly repetitive”

  48. Cot curves eucaryotes show 3 step curves Step 1 renatures rapidly: “highly repetitive” Step 2 is intermediate: “moderately repetitive”

  49. Cot curves eucaryotes show 3 step curves Step 1 renatures rapidly: “highly repetitive” Step 2 is intermediate: “moderately repetitive” Step 3 is ”unique"

  50. Molecular cloning To identify the types of DNA sequences found within each class they must be cloned

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