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Basic Molecular Biology

Basic Molecular Biology. Many slides by Omkar Deshpande. Overview. Structures of biomolecules Central Dogma of Molecular Biology Overview of this course Genome Sequencing. Human Genome Program, U.S. Department of Energy, Genomics and Its Impact on Medicine and Society: A 2001 Primer, 2001.

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Basic Molecular Biology

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  1. Basic Molecular Biology Many slides by Omkar Deshpande

  2. Overview • Structures of biomolecules • Central Dogma of Molecular Biology • Overview of this course • Genome Sequencing

  3. Human Genome Program, U.S. Department of Energy, Genomics and Its Impact on Medicine and Society: A 2001 Primer, 2001

  4. Watson and Crick

  5. Nucleic acids (DNA and RNA) • Form the genetic material of all living organisms. • Found mainly in the nucleus of a cell (hence “nucleic”) • Contain phosphoric acid as a component (hence “acid”) • They are made up of nucleotides.

  6. Nitrogenous Base Nitrogenous Base Phosphate Group Phosphate Group Sugar Sugar Nucleotides

  7. A T G C C G G C A T C G A T G C DNA A = T G = C

  8. The gene and the genome • Genome = The entire DNA sequence within the nucleus. • The information in the genome is used for protein synthesis • A gene is a length of DNA that codes for a (single) protein.

  9. How big are genomes?

  10. Repeats • The DNA is full of repetitive elements (those that occur over & over & over) • There are several type of repeats, including SINEs & LINEs (Short & Long Interspersed Elements) (1 million just ALUs) and low complexity elements. • Their function is poorly understood, but they make problems more difficult.

  11. Central dogma ZOOM IN tRNA transcription DNA rRNA snRNA translation POLYPEPTIDE mRNA

  12. Transcription • The DNA is contained in the nucleus of the cell. • A stretch of it unwinds there, and its message (or sequence) is copied onto a molecule of mRNA. • The mRNA then exits from the cell nucleus.

  13. A A T G G C C C G G G C A A T C C G U A T G G C DNA RNA A = T G = C T  U

  14. More complexity • The RNA message is sometimes “edited”. • Exons are nucleotide segments whose codons will be expressed. • Introns are intervening segments (genetic gibberish) that are snipped out. • Exons are splicedtogether to form mRNA.

  15. Splicing frgjjthissentencehjfmkcontainsjunkelm thissentencecontainsjunk

  16. Key player: RNA polymerase • It is the enzyme that brings about transcription by going down the line, pairing mRNA nucleotides with their DNA counterparts.

  17. Promoters • Promoters are sequences in the DNA just upstream of transcripts that define the sites of initiation. • The role of the promoter is to attract RNA polymerase to the correct start site so transcription can be initiated. 5’ 3’ Promoter

  18. Promoters • Promoters are sequences in the DNA just upstream of transcripts that define the sites of initiation. • The role of the promoter is to attract RNA polymerase to the correct start site so transcription can be initiated. 5’ 3’ Promoter

  19. Transcription – key steps DNA • Initiation • Elongation • Termination DNA + RNA

  20. Transcription – key steps DNA • Initiation • Elongation • Termination

  21. Transcription – key steps DNA • Initiation • Elongation • Termination

  22. Transcription – key steps DNA • Initiation • Elongation • Termination

  23. Transcription – key steps DNA • Initiation • Elongation • Termination DNA + RNA

  24. Genes can be switched on/off • In an adult multicellular organism, there is a wide variety of cell types seen in the adult. eg, muscle, nerve and blood cells. • The different cell types contain the same DNA though. • This differentiation arises because different cell types express different genes. • Promoters are one type of gene regulators

  25. Transcription (recap) • The DNA is contained in the nucleus of the cell. • A stretch of it unwinds there, and its message (or sequence) is copied onto a molecule of mRNA. • The mRNA then exits from the cell nucleus. • Its destination is a molecular workbench in the cytoplasm, a structure called a ribosome.

  26. Translation • How do I interpret the information carried by mRNA to the Ribosome? • Think of the sequence as a sequence of “triplets”. • Think of AUGCCGGGAGUAUAG as AUG-CCG-GGA-GUA-UAG. • Each triplet (codon) maps to an amino acid.

  27. The Genetic Code • f : codon amino acid • 1968 Nobel Prize in medicine – Nirenberg and Khorana • Important – The genetic code is universal! • It is also redundant / degenerate.

  28. The Genetic Code

  29. Proteins • Composed of a chain of amino acids. R | H2N--C--COOH | H 20 possible groups

  30. Proteins R R | | H2N--C--COOH H2N--C--COOH | | H H

  31. Dipeptide This is a peptide bond R O R | II | H2N--C--C--NH--C--COOH | | H H

  32. Protein structure • Linear sequence of amino acids folds to form a complex 3-D structure. • The structure of a protein is intimately connected to its function. The 3-D shape of proteins gives them their working ability – the ability to bind with other molecules.

  33. Our course (2417) Part 1, DNA: Assembly, Evolution, Alignment Part 2, Genes: Prediction, Regulation Part 3, Interactions transcription DNA rRNA snRNA translation POLYPEPTIDE mRNA

  34. DNA Sequencing Some slides shamelessly stolen from Serafim Batzoglou

  35. DNA sequencing How we obtain the sequence of nucleotides of a species …ACGTGACTGAGGACCGTG CGACTGAGACTGACTGGGT CTAGCTAGACTACGTTTTA TATATATATACGTCGTCGT ACTGATGACTAGATTACAG ACTGATTTAGATACCTGAC TGATTTTAAAAAAATATT…

  36. Which representative of the species? Which human? Answer one: Answer two: it doesn’t matter Polymorphism rate: number of letter changes between two different members of a species Humans: ~1/1,000 – 1/10,000 Other organisms have much higher polymorphism rates

  37. Why humans are so similar A small population that interbred reduced the genetic variation Out of Africa ~ 40,000 years ago Out of Africa

  38. Migration of human variation http://info.med.yale.edu/genetics/kkidd/point.html

  39. http://info.med.yale.edu/genetics/kkidd/point.html Migration of human variation

  40. http://info.med.yale.edu/genetics/kkidd/point.html Migration of human variation

  41. DNA Sequencing Goal: Find the complete sequence of A, C, G, T’s in DNA Challenge: There is no machine that takes long DNA as an input, and gives the complete sequence as output Can only sequence ~500 letters at a time

  42. DNA sequencing – vectors DNA Shake DNA fragments Known location (restriction site) Vector Circular genome (bacterium, plasmid) + =

  43. Different types of vectors

  44. DNA sequencing – gel electrophoresis • Start at primer (restriction site) • Grow DNA chain • Include dideoxynucleoside (modified a, c, g, t) • Stops reaction at all possible points • Separate products with length, using gel electrophoresis

  45. Electrophoresis diagrams

  46. Challenging to read answer

  47. Challenging to read answer

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