Bio 99 Final Lec 15-16

  1. Codon
    trinucleotide sequence that codes for an amino acid
  2. Crick predicts existence of an _____ ____ that reads DNA (or mRNA) sequences and “translates” to amino acid sequence
    adapter molecule
  3. Transfer RNA connects _____ and _____
    mRNA; protein
  4. anticodon
    triplet nucleotide sequence on tRNA that base pairs with codon on mRNA
  5. The ____ position of the anticodon binds to the ____ position of the
    codon, and vice versa
    first; third
  6. Anticodons will be listed _' to _'
    3; 5
  7. If the codon on the
    mRNA is GCA (5’ to 3’),
    what is its anticodon?

    C. )
  8. There are only ___ amino acids for ___ amino acid-encoding codons (+ __ stop codons)
    20; 61; 3
  9. The Genetic Code is ________
  10. degenerate
    multiple codons can encode the same amino acid. Consequence of there being 64 possible codons but only 20 amino acids
  11. Codon family
    when 4 codons specify the same amino acid
  12. Start Codon
    AUG- Met (methionine)
  13. Stop codons (3)
    • UAA, UAG, UGA
    • no tRNAs that recognize stop codons
  14. Genetic code is read in .....
    non-overlapping triplets (3 nucleotides at a time)
  15. an mRNA has __# of reading frames depending on which nucleotide you start at
  16. A double stranded DNA can potentially be transcribed in either direction leading to __# of potential of reading frames
  17. open reading frame
    A sequence that has a start codon, then a long stretch of codons, and then a stop codon all in the same reading frame
  18. not a question just a tip...
    How do you tell if a DNA or RNA sequence contains an open reading
    1) Does it have a start codon? (AUG)
    2) Does it have a stop codon? (UAA, UAG, UGA)
    3) Are the start and stop codons in the same reading frame?
  19. Inosine can pair with _, _, or _
    C, U, A
  20. Adenosine in 1st anticodon position
    of tRNA is converted to _____
    inosine (I)
  21. A minimum of ___ tRNAs can potentially decode all codons
  22. Humans have ___ different tRNAs
  23. There are many copies of each tRNA
    To ensure survival if any of them are mutated Also ensures there’s enough of each tRNA being transcribed
  24. The 61 codons can potentially be recognized by ___ different tRNAs, due to wobble
  25. In humans, there are __ tRNA genes, and most anticodons are shared by several copies of the same basic  tRNA gene
  26. Codon Bias
    Some amino acids prefer particular codons, offering additional ways to regulate translation
  27. Mutations happen to ___ (not ____), but affect the _____ sequence
    DNA; RNA; mRNA
  28. Single base substitutions: (define and give the 3 types)
    • Change of a single base in the DNA sequence of a gene
    • 1.) Silent 
    • 2.) Missense
    • 3.) Nonsense
  29. Silent
    • change in codon that does not change the amino acid sequence.
    • Example: GAA (Glu) mutated to GAG (Glu). Both encode Glu so the mutation is silent.
  30. Missense
    • change in codon that results in a different amino acid encoded
    • Example: GAA (Glu) mutated to GAC (Asp). Changes Glu to Asp.
    • Some missense mutations are worse than others. Glu and Asp are both negatively  charged and their substitution likely wouldn’t have a huge impact on the protein (unless in the catalytic site)
  31. Nonsense
    • change in codon that creates an early stop codon
    • Example: GAA (Glu) mutated to TAA (UAA). Causes change from Glu to stop codon. Leads to a truncated protein that is often non-functional
  32. Transition mutation
    A purine is substituted for another purine. A to G or G to A. Most common type of mutation. Can lead to all 3 types of substitutions.
  33. Frameshift mutants
    insertion or deletion of nucleotides that alter the reading frame of the coding sequence
  34. Most frameshift mutations result in .....
    premature stop codon
  35. Deletion mutants
    deletion of one or more nucleotides. Sometimes, large blocks of genetic material are missing.
  36. If the number of nucleotides inserted or deleted is the same, then the
    reading frame is ______
  37. reversion mutation
    A reversion mutation is where a mutation that is deleterious to the function of the encoded protein acquires a second mutation that allows the protein to function again
  38. Examples of reversion mutation
    • 1) A missense mutation (Lys to Gly) in the active site causes the protein to be non functional. A second mutation returns the amino acid to Lysine.
    • 2) Same situation as above, but a second mutation changes the Gly to Arg, which is similar to Lys and the protein is mostly functional.
    • 3) A nonsense mutation (Tyr to STOP) creates a truncated protein that is non functional. A second mutation changes STOP back into Tyr.
    • 4) Same situation as above, but the second mutation changes STOP to Ser, allowing a full length protein and restores function. 
    • 5) A frameshift mutation is caused by the insertion of a single nucleotide. A second mutation causes a deletion of a single nucleotide near the site of the insertion. Protein function is restored.
  39. in order for a reversion mutation to be identified, protein function must be .....
    restored (fully or partially)
  40. What causes single base substitutions?
    Chemical carcinogens and many, many other things.
  41. What causes frame shift mutations?
    • Intercalating agents insert themselves into the DNA double helix
    • Cause stretching of the DNA
    • During DNA replication, as the DNA polymerase passes through strands stretched by intercalating agents, they can cause insertion or deletion mutations
    • Acrididin Orange is an intercalating agent used to make frameshift mutations in DNA
  42. Ionizing Radiation
    • a type of radiation that causes the release of electrons from molecules, ionizing them.
    • Can directly damage DNA
    • Can indirectly damage DNA by creating free radicals from water molecules, which damage DNA
    • Leads to double strand DNA breaks
    • During repair, these breaks can lead to large deletion mutations of several nucleotides in length
    • X-rays are a common form of ionizing radiation used to create mutations
  43. Overview: Single base substitution
    Silent, missense, nonsense mutations (carcinogens)
  44. Overview: Frameshift mutations
    insertion or deletion (intercalating agents)
  45. Overview: Deletion mutations
    loss of large chunk of sequence (ionizing radiation)
  46. Overview: Transition mutation
    A to G or G to A
  47. Overview: Reversion mutations
    second mutation that restores protein function
  48. Crick and Brenner: T4 Bacteriophage
    Define.... T4 Bacetriophage, Plaque Assay, B gene, Acridine, and the actual experiment
    • T4 Bacteriophage – virus that infects bacteria
    • Plaque assay – when mixed with bacteria and plated onto agar dishes, the plate will be confluent with growing bacteria, but not where bacteriophage is growing. The plaques are the dark regions with dead bacteria.
    • B gene – required for bacteriophage to infect multiple strains of E. coli. If mutated, will only grow on one strain.
    • Acridine – intercalating agent, low doses introduce single insertion/deletion mutations
    • Experiment – mutate B gene, try to grow on two E. coli strains. Non-functional B gene will allow it to only grow on one strain.
  49. Crick and Brenner: T4 reversion mutants
    • Two mutant strains of Bacteriophage could be crossed and recombine their genetic material. If one strain had a single insertion, and the other strain a single deletion, then combined the two frameshift mutations would offset each other, and the normal reading frame would be restored. Could classify single mutants into two groups by whether they could complement one another: “+” group and “–” group.
    • Two plus mutants won’t complement one another, and two minus mutants won’t complement one another, but one plus and one minus will complement one another.
  50. First evidence that code is read
    in triplets
    • Crick and Leslie Barnett
    • Recombined three “plus” mutant strains
    • 3 insertion (or deletion) mutations will restore reading frame
  51. Why was it important for Crick to use a mutagen that created insertion/
    A) Single base substitutions would not answer the question about gaps in the
    B) Single base substitutions cannot undergo reversion mutations
    C) Unlike 3 insertions mutations, 3 single base substitutions would not
    necessarily restore the protein function when recombined.
    D) Single base substitutions would not affect the B gene’s functions
  52. What tools do you need to decipher the genetic code?
    • 1) Cell free in vitro translation system
    • Add ribosomes, tRNA, mRNA and produce polypeptides
    • 2) mRNA of defined sequence
    • Needed to know the sequences of the mRNA you were adding
    • How do you stitch individual nucleotides together to get fully defined sequences?
    • 3) Method to determine which polypeptides were produced
    • Needed to know which amino acids were generated
  53. A “cell free” system for synthesizing protein
    Steps: (5)
    • 1. Lyze bacterial cells (“cell extracts”)
    • 2. Destroy endogenous mRNA with endogenous RNases
    • 3. Add DNase to destroy DNA
    • 4. Add synthetic mRNA
    • 5. Add radioactive amino acid (1 radioactive amino acid per tube, 20 tubes)
  54. Marshall Nirenberg and Heinrich Matthaei
    • Created method to create synthetic RNAs
    • Polynucleotide phosphorylase – enzyme that normally degrades RNA, but if given excess NDP, will catalyze the reverse reaction and start attaching them together
    • They couldn’t control the sequence, but if they added only UDP, could create a polymer of poly (U) RNA (i.e. UUUUUUUUUU)
    • Created 4 different polynucleotides:
    • • Poly (U) – UUUUUUUUU
    • • Poly (A) – AAAAAAAAAA
    • • Poly (C) – CCCCCCCCC
    • • Poly (G) – GGGGGGGG
  55. Severo Ochoa
    mixed ADP and CDP at 5:1 ratio, to produce triplets of different frequencies
  56. Gobind Khorana
    created dinucleotide, trinucleotide, and tetranucleotide repeats
  57. Maxine Singer
    • Produced
    • trinucleotides of defined
    • sequence (not long polymers
    • of semi-random sequence)
  58. Niremberg and Leder
    • 1) Trinucleotides (not long random polynucleotides)
    • 2) Ribosomes (cell extracts)
    • 3) tRNA (with radioactive amino acids
    • attached)
    • Ran mixture over nitrocellulose filter
    • Filter traps ribosomes with trinucleotides and correctly matched tRNAs
    • unbound trinucleotides and tRNAs pass through
    • Could identify which amino acid/tRNA matched the trinucleotide based on radiolabel
  59. Ribosome filter binding assay
    A single trinucleotide would cause one and only one aminoacyl-tRNA to bind to the ribosome. 61 out of 64 possible codons could be decoded this way
Card Set
Bio 99 Final Lec 15-16