LIFESCI 4 Lecture 12: LIFESCI 4 - Lecture 12 - Complementation and Recombination Analysis Continued

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20 May 2018
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LIFESCI 4 - Lecture 12 - Complementation and Recombination Analysis Continued
Crick et al. studied revertants of rII mutants
- Frameshift mutations
- Answered how the genetic code worked
- Triplet codon is non-overlapping
- A triplet set of codon codes for an amino acid
- Proflavin: a mutagen that causes frameshift mutation
rII mutant (FC0)
- A ๎€frameshift insertion๎€ โ†’ the whole reading frame shifts due to the insert
- โ†’ have wild type phenotypeII (F C0 revertants)r+
- 2 possibilities:
1) True revertant๎€: the exact original insertion was taken out โ†’ sequence
returns to original
2) New mutation (suppressor):๎€ a deletion occurs at a different site that
brings the reading frame back to place, but the original insertion is still
present
Reverting Frameshift Mutants
- 5โ€™ is the N-terminus where 3โ€™ is the C-terminus
- Synthesized from 5โ€™ to 3โ€™
- For the following example sequence, SER - ALA - THR needs to be expressed
[provided by Professor Pham]
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- Line 1 = original sequence
- Line 2 = frameshift insertion at (Arg) โ†’ insertion of Guanine causes frameshift
mutation โ†’ SER - ALA - THR is not expressed
- Line 3 = reverting occurs (suppression) โ†’ after Guanine was inserted, Thymine
at (Ala) was removed and the frame is reset to normal โ†’ SER - ALA - THR is
expressed
Predict the Phenotypes
- If FC0 is insertion, everything else are suppressors (FC1, FC10, FC11)
- Suppressors means that they are deletions (to compensate for the insertion
mutation)
-Double mutation
๎€
: there are 2 mutations โ†’ one insertion + one deletion etc.
-Triple mutation
๎€
: there are 3 mutations โ†’ one insertion + two deletions etc.
Genetic Code
-Start๎€ codon โ†’ ๎€AUG = met
๎€
(this counts as an amino acid)
-Stop๎€ codons โ†’ ๎€UAA, UAG, UGA
๎€
(these are not considered amino acids)
rII Locus Recombination / Complementation Test
- Recombination test is done to find the map distance between the mutants
- If there are no complementation, there should be 0 phages coming out on the K plate
(since it is non-permissive)
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Document Summary

Lifesci 4 - lecture 12 - complementation and recombination analysis continued. Crick et al. studied revertants of rii mutants. A triplet set of codon codes for an amino acid. Proflavin: a mutagen that causes frameshift mutation rii mutant (fc0) A frameshift insertion the whole reading frame shifts due to the insert. 5" is the n-terminus where 3" is the c-terminus. For the following example sequence, ser - ala - thr needs to be expressed. Line 2 = frameshift insertion at (arg) insertion of guanine causes frameshift mutation ser - ala - thr is not expressed. Line 3 = reverting occurs (suppression) after guanine was inserted, thymine at (ala) was removed and the frame is reset to normal ser - ala - thr is expressed. If fc0 is insertion, everything else are suppressors (fc1, fc10, fc11) Suppressors means that they are deletions (to compensate for the insertion mutation) Double mutation : there are 2 mutations one insertion + one deletion etc.

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