Year Morgan and fellow researchers found that chromosomes contained DNA, RNA, and protein.

Transcription

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5 Year 1920 Morgan and fellow researchers found that chromosomes contained DNA, RNA, and protein. Which one actually carries the genetic information? The stuff that gets passed on from generation to generation?

6 Bacterial Transformations Griffith s Experiment studied Streptococcus pneumoniae, the bacteria responsible for pneumonia S-strain (virulent) - Has protective capsule R-strain (non-virulent) - No capsule

7 Bacterial Transformations Round 1 experiment: test effects of both strains of S. pneumoniae Mice treated with S-strain dead! Mice treated with R-strain alive!

8 Bacterial Transformations Took the S-strain and heated a sample over a flame to kill the S-strain bacterial cells

9 Bacterial Transformations Injected heat-killed S-strain into mouse mouse lived

10 Bacterial Transformations Next, mixed live R-strain with heat-killed S- strain cells then injected it into mouse Something from the dead cells had entered the live non-virulent (R-strain)

11 Bacterial Transformations Something from the dead cells had entered the live non-virulent (R-strain)

12 Bacterial Transformations Avery, MacLeod, McCarty continue with Griffith s experiments (minus the mice) 9 hours later

13 Bacterial Transformations Team Avery took parts of the dead virulent cells and separated their components

14 Bacterial Transformations Results R-strain + carbohydrates nothing happened R-strain + lipid nothing happened R-strain + protein nothing happened R-strain + DNA transformed cells R-strain + RNA nothing happened DNA must be the genetic information!

15 Bacteriophages ( phages ) Attack, kill bacteria

16 Bacteriophages Hershey, Chase Experiment Parts of phage were radioactively labeled DNA labeled with Phosphorous-32 Protein coat labeled with Sulfur-35

17 Bacteriophages

18 DNA content: body vs. sex cells Mitosis each body cell should contain the same amount of the genetic material BUT reproductive cells (sex cells) MUST contain half the amount in body cells analyzing the two types of cells shows that there s more DNA in body cells than sex cells THEREFORE DNA is the genetic material

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22 DNA structure Watson, Crick (and Franklin)

23 DNA structure

24 DNA structure Overall structure is the twisted ladder called a double helix!

25 DNA structure Composed of nucleotides Each nucleotide consists of: Phosphate group Sugar (ribose or deoxyribose) Nitrogen-containing base

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28 Small base Large base

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30 DNA structure Chargaff # of A = # of T # of G = # of C Base-pairing rules A Train Goes Choo-choo A with T G with C

31 DNA reproduction (replication) How do we get the same number of DNA in the cells? REPLICATION! How does it replicate? Here are some theories: Conservative: 2 DNA strands don t separate Semiconservative: 2 DNA strands do separate Dispersive: 2 DNA strands break into pieces

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33 Meselsen-Stahl experiment

34 Meselsen-Stahl experiment

35 Experiments performed by Meselson and Stahl Supported the semiconservative model of DNA replication EXPERIMENT Matthew Meselson and Franklin Stahl cultured E. coli bacteria for several generations on a medium containing nucleotide precursors labeled with a heavy isotope of nitrogen, 15 N. The bacteria incorporated the heavy nitrogen into their DNA. The scientists then transferred the bacteria to a medium with only 14 N, the lighter, more common isotope of nitrogen. Any new DNA that the bacteria synthesized would be lighter than the parental DNA made in the 15 N medium. Meselson and Stahl could distinguish DNA of different densities by centrifuging DNA extracted from the bacteria. 1 RESULTS Bacteria cultured in medium containing 15 N 2 Bacteria transferred to medium containing 14 N Figure DNA sample 4 DNA sample centrifuged centrifuged after 20 min after 40 min (after first (after second replication) replication) Less dense More dense The bands in these two centrifuge tubes represent the results of centrifuging two DNA samples from the flask in step 2, one sample taken after 20 minutes and one after 40 minutes.

36 CONCLUSION Meselson and Stahl concluded that DNA replication follows the semiconservative model by comparing their result to the results predicted by each of the three models in Figure The first replication in the 14 N medium produced a band of hybrid ( 15 N 14 N) DNA. This result eliminated the conservative model. A second replication produced both light and hybrid DNA, a result that eliminated the dispersive model and supported the semiconservative model. First replication Second replication Conservative model Semiconservative model Dispersive model

37 Replication proteins

38 Process of Replication Click here to see an animation

39 Prokaryotic Chromosome

40 Eukaryotic Chromosome

41 From Genes to Protein A.P. Biology

42 Typical Gene Structure Regulatory sites Promoter (RNA polymerase binding site) DNA strand Start transcription Stop transcription

43 RNA Contains ribose instead of deoxyribose Single stranded Nucleotide uracil replaces thymine

44 3 Types of RNA Messenger RNA = mrna -Carries genetic code to the ribosome Transfer RNA = trna -Carries amino acid to the codon on mrna Ribosomal RNA = rrna -forms parts of ribosome

45 Basic Principles of Transcription and Translation Transcription Is the synthesis of RNA under the direction of DNA Produces messenger RNA (mrna) Translation Is the actual synthesis of a polypeptide, which occurs under the direction of mrna Occurs on ribosomes

46 In prokaryotes Transcription and translation occur together TRANSCRIPTION DNA mrna Ribosome TRANSLATION Polypeptide (a) Prokaryotic cell. In a cell lacking a nucleus, mrna produced by transcription is immediately translated without additional processing.

47 In eukaryotes RNA transcripts are modified before becoming true mrna Nuclear envelope TRANSCRIPTION DNA RNA PROCESSING Pre-mRNA mrna Ribosome Figure 17.3b TRANSLATION Polypeptide (b) Eukaryotic cell. The nucleus provides a separate compartment for transcription. The original RNA transcript, called pre-mrna, is processed in various ways before leaving the nucleus as mrna.

48 During transcription The gene determines the sequence of bases along the length of an mrna molecule DNA molecule Gene 1 Gene 2 DNA strand (template) 3 Gene 3 A C C A A A C C G A G T 5 The side of the DNA strand that is used as a template for mrna synthesis is called the cdna (complementary or copy DNA) TRANSCRIPTION mrna 5 TRANSLATION U G G U U U G G C U C A Codon 3 Protein Trp Phe Gly Ser Amino acid

49 In laboratory experiments Genes can be transcribed and translated after being transplanted from one species to another

50 Molecular Components of Transcription RNA synthesis Is catalyzed by RNA polymerase, which pries the DNA strands apart and hooks together the RNA nucleotides Follows the same base-pairing rules as DNA, except that in RNA, uracil substitutes for thymine

51 Synthesis of an RNA Transcript The stages of transcription are Initiation Elongation Termination 5 3 Promoter RNA polymerase 5 3 Unwound DNA 5 3 Rewound RNA 5 RNA RNA Transcription unit Start point transcript transcript DNA Template strand of DNA Initiation. After RNA polymerase binds to the promoter, the DNA strands unwind, and the polymerase initiates RNA synthesis at the start point on the template strand Elongation. The polymerase moves downstream, unwinding the DNA and elongating the RNA transcript 5 3. In the wake of transcription, the DNA strands re-form a double helix Termination. Eventually, the RNA transcript is released, and the polymerase detaches from the DNA Figure Completed RNA transcript 3

52 Elongation Non-template strand of DNA RNA nucleotides RNA polymerase 3 A T C C A A 3 end U 5 A E G C A T A G G T T 5 Direction of transcription ( downstream ) Template strand of DNA Newly made RNA

53 RNA Polymerase Binding and Initiation of Transcription TRANSCRIPTION DNA 1 Eukaryotic promoters RNA PROCESSING Pre-mRNA Promoters signal the initiation of RNA synthesis Transcription factors 5 3 TRANSLATION mrna Ribosome Polypeptide Promoter T A T A A A A A T A T T T T TATA box Start point Template DNA strand 2 Several transcription factors 3 5 Help eukaryotic RNA polymerase recognize promoter sequences 5 3 Transcription factors 3 Additional transcription factors 3 5 RNA polymerase II Transcription factors TATA box Figure 17.8 Transcription initiation complex RNA transcript

54 Transcription Making a Copy of the DNA 1. RNA polymerase separates the DNA strands at a promoter region on the DNA (TATA box) 2. RNA polymerase adds nucleotides in sequence to mrna 3. RNA polymerase falls off the DNA at a terminator sequence on the DNA

55 Unwinding of the DNA Helix by RNA Polymerase Step #1

56 Transcription of mrna Complementary to DNA Step #2

57 Alteration of mrna Ends Each end of a pre-mrna molecule is modified The 5 end receives a modified nucleotide cap The 3 end gets a poly-a tail A modified guanine nucleotide added to the 5 end 50 to 250 adenine nucleotides added to the 3 end TRANSCRIPTION DNA RNA PROCESSING mrna Pre-mRNA G P P P 5 Protein-coding segment Polyadenylation signal AAUAAA 3 AAA AAA TRANSLATION Ribosome Polypeptide 5 Cap 5 UTR Start codon Stop codon 3 UTR Poly-A tail

58 RNA splicing Removes introns and joins exons Ribozymes Are catalytic RNA molecules that function as enzymes and can splice RNA TRANSCRIPTION RNA PROCESSING mrna TRANSLATION DNA Pre-mRNA Ribosome Pre-mRNA 5 5 Cap 1 Exon Intron Coding segment Exon Intron Exon Introns cut out and exons spliced together Poly-A tail Polypeptide mrna 5 Cap Poly-A tail 3 UTR UTR Introns - Intervening sequence = Junk DNA Exons Expressed sequence

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60 mrna Processing in Eukaryotic Cells

61 See handout to review transcription

62 The Genetic Code Order of bases must somehow control order of amino acids in a protein how? DNA A-T T-A G-C C-G C-C A-T controls How? Protein Ala Gly Tyr Ser Phe

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64 The Genetic Code After Francis Crick first described the central dogma, scientists immediately began speculating about how the genetic code worked. Could each letter (base) in the RNA language code for one letter (amino acid) in the protein language? If each of the four bases in mrna coded for one amino acid in a protein, there could only be four different amino acids.

65 The Genetic Code Would it be possible for two-letter combinations, such as AG, to code for amino acids? If you arrange the four bases in pairs, how many different combinations would you have? Arranging the four bases in pairs produces 16 different combinations not quite enough to specify 20 amino acids.

66 The Genetic Code Would three-letter combinations work? The next step: Figuring out which three-letter combinations, called codons, specified each of the 20 amino acids. How would you try to solve the genetic code?

67 Nirenberg & Matthaei Made artificial RNA: polyribonucleotides can be used to make proteins First was polyuridylic acid made protein that consisted of ONLY phenylalanine (Phe) UUU = Phe

68 The Genetic Code Eventually, other codons were worked out AUG = Met (methionine); the start codon UAA, UAG, UGA was the termination or stop codon

69 Notice that all 2 or more codons yields the same amino acid this means that the code is degenerate. Proteins that are manufactured are not always enzymes

70 Transfer RNA (trna) Made by txn (transcription) from trna genes in DNA Has a cloverleaf shape 20 different trna for the 20 different amino acids

71 TRANSCRIPTION DNA Molecules of trna are not all identical TRANSLATION mrna Ribosome Polypeptide Polypeptide Amino acids Each carries a specific amino acid on one end Each has an anticodon on the other end Ribosome trna with amino acid attached Gly trna A A A U G G U U U G G C Anticodon 5 mrna Codons 3

72 acylation occurs here (attachment of a specific amino acid; requires energy) Anticodon= 3 base code that fits onto mrna

73 5 3 Amino acid attachment site Hydrogen bonds A A G Anticodon (b) Three-dimensional structure 3 5 Anticodon (c) Symbol used in this book

74 A specific enzyme called an aminoacyl-trna synthetase Joins each amino acid to the correct trna

75 Breaking the Code Refer to handout

76 The ribosome Is part of the cellular machinery for translation, polypeptide synthesis Facilitates the specific coupling of trna anticodons with mrna codons during protein synthesis

77 Ribosomal RNA (rrna) Consists of 2 subunits: 1 large, 1 small Contains rrna & proteins

78 Protein Synthesis: Translation 1. mrna transcribed from structural genes in DNA Structural genes are genes that code for proteins 2. mrna attaches to small subunit, then to large subunit 3. AUG codon of mrna attaches to P site of large subunit

79 Protein Synthesis: Translation 4. trna with anticodon UAC enters, carrying Met or fmet) and attaches to AUG codon 5. Second trna carries another amino acid to the A site 6. Peptidyl transferase carries Met over & attaches to the second amino acid at A site

80 Protein Synthesis: Translation 7. Both trna molecules are translocated toward 5 end of mrna & original trna (that was carrying Met) is released) 8. A third trna attaches to A site; first 2 amino acids carried over to third amino acid Translocation & peptide building go on until stop codon is reached

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84 Gene Mutations: Substitution, Insertion, & Deletion Substitution Insertion Deletion Point Mutation Frameshift Mutation

85 Mutations Changes in the genetic material Point (substitution) mutations: one nucleotide is substituted for another

86 Mutations Some point mutations may not make a difference in the resulting polypeptide chain because of the degenerate code those are considered silent mutations

87 Mutations Frameshift mutations Insertions or deletions of one or more bases May cause serious changes in the amino acid sequence

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89 Base Pair Substitution Missense mutation altered codon codes for a different amino acid Nonsense mutation altered codon changed to a stop codon (no amino acid delivered)

90 Chromosomal Mutations Deletion Duplication Inversion Translocation

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92 Central Dogma in Biology