1 MSH2 MSH6 Template strand 5' A 3' Newly synthesized daughter strand MLH1 endonuclease, PMS2 2 DNA helicase DNA exonuclease 5' 3' 5' 3' Gap repair by DNA polymerase and ligase 3 5' 3' A |IT||| 3' 5' 31 Lodish et al., Molecular Cell Biology, 9e, © 2021 W. H. Freeman and Company 31

Covalent modification of the E. coli origin of replication (oriC) is required for initiation to proceed. What modification must be made, what is the target of the modification and what enzyme is involved?

Describe a typical program used for DNA amplification with the thermal cycler and the purpose of each steps.

In E. coli, the leading strand is primarily synthesized by:       DNA polymerase ε   DNA polymerase I   DNA polymerase II   DNA polymerase δ   DNA polymerase III

Explain why Mismatch Repair of absolutely requires semi-conservative DNA replication during S phase.

DNA polymerases are capable of editing and error correction, meaning it is able to edit and correct single base error so that the gene is not affected. However, RNA polymerase has a limited capacity for error correction. Given that a single base error in either replication or transcription can lead to error in protein synthesis, suggest a brief explanation for this difference in the capability of error correction between DNA polymerase and RNA polymerase.

DNA polymerase occasionally incorporates the wrong nucleotide during DNA replication. If left unrepaired, the base-pair mismatch that results will lead to mutation in the next replication. As part of a template strand, the incorporated wrong base will direct the incorporation of a base complementary to itself, so the bases on both strands of the DNA at that position will now be different from what they were before the mismatch event. The MER-minus strain of yeast does not have a functional  mismatch excision repair system, but it has normal base excision repair and nucleotide excision repair systems. Which of the following statements is correct about differences in the mutation spectrum between MER-minus and wildtype yeast? More than one answer is correct. Options:   More point mutations will arise in MER-minus yeast.   Fewer point mutations will arise in MER-minus yeast as compared with wildtype.   Of the total point mutations that…

A conditional mutation expresses its mutant phenotype only under certain conditions (the restrictive conditions) and expresses the normal phenotype under other conditions (the permissive conditions). One type of conditional mutation is a temperature-sensitive mutation, which expresses the mutant phenotype only at certain temperatures. Strains of E. coli have been isolated that contain temperature-sensitive mutations in genes encoding different components of the replication machinery. In each of these strains, the protein produced by the mutated gene is nonfunctional under the restrictive conditions. You grow these strains under the permissive conditions and then abruptly switch them to the restrictive conditions. After one round of replication under therestrictive conditions, you isolate DNA from each strain and analyze it. What characteristics would you expect to see in the DNA isolated from a strain with a temperature-sensitive mutation in the gene that encodes the following…

1 (a) What do ApR, TcR, and ori on the pBR322 map represent and discuss there individual functions? (b)Does the undigested plasmid show more than a single band when electrophoresed? Why? (c)What other kinds of molecules, in addition to plasmid DNA would you expect to find in a sample of plasmid DNA extraction? (This is not a graded assignment)

What type of repair would be used to fix a mutation that arises during replication and that escapes detection by polymerase proofreading mechanisms? What is the most likely reason that this mutation escaped correction during proofreading?