Term
| What is the first challenge in gene targeting? |
|
Definition
| Production of a specific mutation in cultured cells |
|
|
Term
| What is the second challenge in gene targeting? |
|
Definition
| How to transfer a specific mutation into the germ line |
|
|
Term
Which challenge did Capecchi work on
? |
|
Definition
| The first; production of a specific mutation in cultured cells |
|
|
Term
Which challenge did Evans work on
? |
|
Definition
| The second; how to transfer a specific mutation into the germ line |
|
|
Term
| What was one of Capecchi's first methodological successes? |
|
Definition
|
|
Term
| Why was the discovery that the insertion of targeted DNA was governed by homologous recombination important? |
|
Definition
| It showed that mammalian cells have machinery to integrate DNA in a specific order, at specific locations |
|
|
Term
| True or False: HR is cell-cycle dependent |
|
Definition
|
|
Term
| which is more efficient, microinjection or electroporation? |
|
Definition
|
|
Term
| if microinjection is more efficient, why did Capecchi choose electroporation? |
|
Definition
| You can use a lot more cells at a single time |
|
|
Term
| G418 is analogous to what antibiotic |
|
Definition
|
|
Term
| True or false: all DNA is recombinant DNA? |
|
Definition
|
|
Term
| When does the majority of DNA recombination occur? |
|
Definition
|
|
Term
| When is the only time chromosomal DNA order changes? |
|
Definition
| When transposons move around |
|
|
Term
|
Definition
| genetic variation, retrieve lost sequences, restart stalled/damaged repln forks, regulation of gene expression |
|
|
Term
| What techniques are available due to an understanding of HR? |
|
Definition
| knockout, transgenic orgs |
|
|
Term
|
Definition
| when ss region of DNA pairs with complementary strand of the homologue |
|
|
Term
| What are the underlying mechanisms of branch migration? |
|
Definition
| melting a parental bp while forming a bp in the new strand |
|
|
Term
| What is the one drawback to the Holliday model? |
|
Definition
| It does not mention new DNA synth at all |
|
|
Term
|
Definition
| when the base pairs don't match up during HR |
|
|
Term
|
Definition
| when the DNA is "peeled" away from its complementary strand so it can bind across the homologue |
|
|
Term
| What is the relationship between the cut strands during HR cleavage? |
|
Definition
| The 2 strands with the same sequence and polarity must be cleaved |
|
|
Term
| which strands are cut with site 1 cleavage? |
|
Definition
| the unbroken strands; those not involved in the Holliday junction |
|
|
Term
| True or false: the patch product leads to reassortment of genes |
|
Definition
|
|
Term
| Why is DSB a better model for HR? |
|
Definition
| DSB's occur frequently under normal conditions, as opposed to ss nicks |
|
|
Term
| what is an alternative to HR? |
|
Definition
| non-homologous end joining |
|
|
Term
| What is the main reason for HR in bacteria? |
|
Definition
|
|
Term
| What are two causes of stalled replication fork? |
|
Definition
| ss nick, lesion (distortion due to mismatch) |
|
|
Term
| What is the difference between recombination and homologous recombination? |
|
Definition
| HR for repair, recomb during meiosis |
|
|
Term
| What is the major role of RecBCD? |
|
Definition
| process DSB's to form ss tails |
|
|
Term
| What enzyme does RecBCD recruit? |
|
Definition
|
|
Term
| What is the notation for a gene? |
|
Definition
|
|
Term
| T/F: RecBCD is ATP-dependent |
|
Definition
|
|
Term
|
Definition
| crossover hotspot initiation |
|
|
Term
| Which subunit is the nuclease activity associated with? |
|
Definition
| none, it is a characteristic of the combined properties of RecB and RecD |
|
|
Term
| True or False: RecB and RecD move at the same speed? |
|
Definition
| False, an ss loop RecB strand |
|
|
Term
| Which direction does RecB move? |
|
Definition
|
|
Term
| Which subunit is typically inactivated after RecBCD reaches the chi site? |
|
Definition
|
|
Term
| What is the expected chi site frequency in E coli? |
|
Definition
| ~ every 65 bp (80 times total) |
|
|
Term
| What is the observed chi site frequency in E coli? |
|
Definition
| 1009 times in the genome (as opposed to 80) |
|
|
Term
| What is the central protein in HR? |
|
Definition
|
|
Term
| name the substrate for RecA |
|
Definition
|
|
Term
| What are the 3 requirements for RecA? |
|
Definition
1. complementary seq
2. ssDNA so RecA can bind
3. DNA end where the two strands can intertwine |
|
|
Term
| Describe the general structure of RecA |
|
Definition
|
|
Term
| How does the structure of RecA accommodate DNA sampling? |
|
Definition
| the DNA becomes extended (larger distances between bp) |
|
|
Term
|
Definition
| Can do both, prefers 3' to 5' |
|
|
Term
| What is the proposed mechanism for DNA sampling in the secondary site of RecA? |
|
Definition
|
|
Term
| True/False: All organisms have RecA analogs |
|
Definition
|
|
Term
| What is /are the euk RecA analogs? |
|
Definition
|
|
Term
| What is the major role of RuvAB? |
|
Definition
| Holliday junction migration |
|
|
Term
| What is the major role of RuvA? |
|
Definition
| recognizes structure of Holliday jxn, recruits RuvB |
|
|
Term
| Describe the functions of RuvC |
|
Definition
| Complexes with RuvA and RuvB, recogs Holl jxn, nicks strand with same polarity |
|
|
Term
| Describe junction recognition by RuvC |
|
Definition
| RuvC recognizes the structure, not the sequence |
|
|
Term
| What is the consensus sequence for RuvC? |
|
Definition
|
|
Term
| What is useful about RuvC's modest selectivity? |
|
Definition
| It allows for a region of branch migration to occur before cleavage |
|
|
Term
| Define meiotic recombination |
|
Definition
|
|
Term
| T/F Spo11 is sequence dependent |
|
Definition
| False, it is time dependent. Acts during meiosis |
|
|
Term
| What is the role of Spo11? |
|
Definition
|
|
Term
| T/F: Spo11 cuts with high sequence specificity |
|
Definition
| False, low specificity, but at very precise times |
|
|
Term
| What is Spo11's preferred cut site? |
|
Definition
| At sites with low nucleosome count (spatial reasons) |
|
|
Term
| What is the mechanism of action for Spo11? |
|
Definition
| A tyrosine side chain attacks the phosphodiester backbone of DNA |
|
|
Term
| Which protein is responsible for ss processing in euk? |
|
Definition
|
|
Term
| Describe the structure of MRX |
|
Definition
| Complex of 3 different proteins |
|
|
Term
| What is the prokaryotic analog of MRX? |
|
Definition
|
|
Term
| T/F MRX and Spo11 act in tandem |
|
Definition
|
|
Term
| In which direction does MRX work? |
|
Definition
|
|
Term
| What are the two euk RecA anologs? |
|
Definition
|
|
Term
| Recombination occurs between which members of the homologue? |
|
Definition
|
|
Term
| What is the role of Rad52? |
|
Definition
| Promotes assembly of Rad51-DNA filaments |
|
|
Term
| Describe the action of Rad52 |
|
Definition
| Antagonizes RPA so that Rad51 can bind instead |
|
|
Term
|
Definition
| ss binding protein in euk |
|
|
Term
| What genes are present in a cells? |
|
Definition
|
|
Term
| What genes are present in alpha cells? |
|
Definition
|
|
Term
| What is the role of HO endonuclease? |
|
Definition
| intro DSB during mating type switching |
|
|
Term
|
Definition
| synthesis-dependent strand annealing; a new recombination model to explain gene conversion without crossing over (eg mating type switching) |
|
|
Term
| Describe the replication fork in the SDSA model |
|
Definition
| a full replication fork, with leading and lagging strands |
|
|
Term
| Describe the two methods of obtaining a gene conversion? |
|
Definition
| mating type switching and mismatch repair |
|
|
Term
| Mnemonic for DNA/RNA stop codons |
|
Definition
| u are away, u go away, u are gone=> UAA, UGA, UAG and TAA, TGA, TAG |
|
|
