Biochemii Lect 14,15,16

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Are Ter DNA polymerase traps directional?

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Yes

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Eukaryotic DNA polymerase delta is _____ slower than prokaryotic DNA pol3.

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20x

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In eukaryotic DNA replication termination RNA primer degradation on the __________ strand results in a terminal gap and a ______ overhang.

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lagging, 3'

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Each round of replication for a linear chromosome reults in?

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a shortening of the strands

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The solution for dealing with the ends of linear chromosome shortening is?

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adding a set of repeated sequences, non-encoding that can be copied using a special enzyme (telomeres)

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What enzyme adds nucleotide bp's to a linear chromosome?

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Telomerase

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Tell me about this.

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Yep you are right. GOOD JOB!!

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What is the human telomere sequence?

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TTAGGG repeated approx 1,500 times

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In humans how many DNA lesions per person per second?

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10,000,000,000,000

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According to Lehninger what is the ratio of DNA lesions to mutations?

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less than 1 in 10000

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Ultimately what does a DNA mutation on an exon lead to?

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alterations in a protein

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If a basepair mutation (change) results in a change to an important amino acid for protein function is causes?

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a deleterious mutation, often a disease state

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A mutation is defined as a ____________ in the base sequence of ________.

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heritable change, DNA

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Define a silent mutation?

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affects nonessential DNA or has a negligible effect on gene function

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Define a nonsilent mutation?

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They are deleterious of neutral, they are detrimental or confer no biological advantage to the affected organism

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If a basepair change results in a change to an important amino acid for protein function it usually causes?

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deleterious mutations and often a disease state.

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What kind of bacterial cells are used in the Ames test?

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mutants that are unable to produce his

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What kind of medium is used in the Ames test?

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His free medium

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What is a putative mutagen?

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putative = potential or possible

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Describe the Ames test and how to analyze it?

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The Ames test is a test for?

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measuring of mutagenicity

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There is a strong correlation between mutagenesis and?

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cancer

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Are all cancers mutagenic?

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no

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Draw a graph of the Ames test. Concentration of mutagen vs. distance from the center of disk.
Name and label the 2 zones we should know.

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Name the 5 repair systems we should know.

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Methyl-directed Mismatch Repair (MMR)
Base Excision Repair (BER)
Nucleotide Excision Repair (NER)
Direct Repair
Recombination Repair

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MMR is used to correct _______ mismatches that occur during DNA _________.

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rare, replication

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MMR requires what two things to function?

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intact template (old) strand
ability to discriminate between the old and new strands (needs a hemimethylated strand?

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What is the base methylated base sequence that MMR recognizes?

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GATC methylated a N6 of the adeninine

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Which mut proteins form complexes that attach to mismatched DNA?

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mut S
mut L

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DNA is __________ through the mutL-mutS complex until it finds ______________________________.

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looped, methyl group on GATC

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In the MMR repair system the distance between the mismatch and the nearest methylated adenine can be up to _________bp.

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1000

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Which protein in the MMR repair system acts a an endonuclease and nicks the nascent DNA at the methylated site of the parent strand?

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mut H

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In MMR repair the methyl group on the parent strand activates an _______________ activity in mutH.

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endonuclease

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The endoncuclease activity of mutH cuts the nascent strand on the ___ end of the GATC sequence.

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5'

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After mutH creates a nick in the nascent strand what happens?

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the appropriate exonuclease cuts out the bp back to the mismatch, then poly3 comes in and fixes the gap

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Which mut protein scans the strands for a mismatch in MMR?

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mut S

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The MMR repair process is energetically expensive. Why?

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up to 1000 bases are removed and replaced to correct for one mismatch

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The BER replaces base lesion due to?

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DNA glycoslyases in the BER system hydrolyze the _____________ bond.

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N-glycosidic bond

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DNA glycoslyases in the BER system are ____________ for a given base.

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specific

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DNA glycoslyases in the BER system are specific for _________ and ____________.

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a given base, also specific to damaged type of bases

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What does MMR, BER, and NER stand for in DNA repair systems?

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methyl-directed mismatch repair
base excision repair
nucleotide excision repair

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NER is used to repair ___________ lesions that cause _____________ changes in DNA.

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bulky, conformational

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What are two kinds of bulky lesions?

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Name two causes of base adducts.

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What is unique about the nuclease activity of NER?

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Mutations in the exinuclease of the homologous human pathway of NER can cause?

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MMR uses DNA poly _____, while NER and BER uses DNA poly ____.

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3,1

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In the BER repair system a damaged base is recognized by a ______________________.

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DNA glycosylase

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Direct nucleotide repair involves direct repair of a _________ base without ___________ if the phosphodiester backbone.

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defective, cleavage

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Direct nucleotide repair involves direct repair of a defective base without cleavage of the _____________ ____________.

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phosphodiester backbone

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Some examples of direct nucleotide repair.

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Some examples of direct nucleotide repair.

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Some examples of direct nucleotide repair.

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Some examples of direct nucleotide repair.

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Some examples of direct nucleotide repair.

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Some examples of direct nucleotide repair.

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In the event of an O6-methylated guanine the first round of replication will produces?

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G-T

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O6-methylguanine is corrected in the direct nucleotide repair system by_____________________________.

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This repair mechanism is in which repair system? This reaction is not __________, and not ___________.

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direct nucleotide repair, catalytic, reversible

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This direct nucleotide repair mechanism is considered energetically expensive because?

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The recombination repair method is different from MMR, NER, and BER because?

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it does not require a template

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If an O6-methylguanine is left uncorrected in DNA it will convert a ____ pair to a _____ pair.

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G-C to A-T

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Are double strand breaks (DBS) common in DNA?

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yes

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the __________ into a gap by the RecBCD complex
2. Strand invasion via the recA protein complex
3. 3' end extension; branch migration
4. 3' end extension
5. resolution of Holliday intermediate by RuvC

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DSB (double strand break)

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the ___________ complex
2. Strand invasion via the recA protein complex
3. 3' end extension; branch migration
4. 3' end extension
5. resolution of Holliday intermediate by RuvC

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recBCD

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the RecBCD complex
2. via the recA protein complex
3. 3' end extension; branch migration
4. 3' end extension
5. resolution of Holliday intermediate by RuvC

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Strand invasion

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the RecBCD complex
2. Strand invasion via the ____________ complex
3. 3' end extension; branch migration
4. 3' end extension
5. resolution of Holliday intermediate by RuvC

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recA protein

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the RecBCD complex
2. Strand invasion via the recA protein complex
3. ____________; branch migration
4. 3' end extension
5. resolution of Holliday intermediate by RuvC

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3' end extension

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the RecBCD complex
2. Strand invasion via the recA protein complex
3. 3' end extension; branch migration
4. ______________
5. resolution of Holliday intermediate by RuvC

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3' end extension

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the RecBCD complex
2. Strand invasion via the recA protein complex
3. 3' end extension; branch migration
4. 3' end extension
5. resolution of __________ intermediate by RuvC

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Holliday

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the RecBCD complex
2. Strand invasion via the recA protein complex
3. 3' end extension; branch migration
4. 3' end extension
5. resolution of Holliday intermediate by ________

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ruvC

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This is called?

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D-loop

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This is called?

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double holliday junction or intermediate

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5 STEPS IN DNA RECOMBINATION REPAIR:
1. Expanding the DSB (double strand break) into a gap by the RecBCD complex
2. Strand invasion via the recA protein complex
3. 3' end extension; ___________________
4. 3' end extension
5. resolution of Holliday intermediate by RuvC

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branch migration

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In DNA recombination the _____________ has both helicase and nuclease activities.

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RecBCD complex

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In DNA recombination the RecBCD complex has both __________ and _________ activities.

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helicase, nuclease

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In DNA recombination the RecBCD complex initiates DSB repair by attaching to and degrading free ________________.

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double strand ends

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In DNA recombination the ________ complex initiates DSB repair by attaching to and degrading free double strand ends.

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RecBCD

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In DNA recombination the RecBCD complex initiates DSB repair by ________ to and _________ free double strand ends.

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attaching, degrading

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What is the purpose of the chi-scanning site in the RecBCD complex?

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enzyme binding to the chi sequence halts degradation of the 3'end while degradation of the 5'end continues

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In DNA recombination repair what step is the RecBCD complex involved in?

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1. expanding the DSB into a gap

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Does more bp per turn mean overwinding or underwinding?

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underwinding

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The RecA protein complex is involved in what step of DNA recombination repair?

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2. strand invasion

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The RecA protein polymer form ______________ helical polymer filaments that fit into the ________ groove of DNA molecules.

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right-handed, major

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The RecA protein polymer assembles on an existing ________ in the 5'--3' direction.

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ssDNA

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The RecA protein polymer assembles on an existing ssDNA in the _______ direction.

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5'--3'

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RecA can facilitate ______________ and _____________.

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strand invasion, branch migration

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The RecA protein polymer can mediate strand invasion only if presented with _________.

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ssDNA

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In DNA recombination repair what protein complex is associated with step 1: expanding the DSB into a gap.

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RecBCD

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In DNA recombination repair what protein complex is associated with step 2: strand invasion?

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RecA

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In DNA recombination repair what enzymes are associated with step 3: 3' end extension branch migration?

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DNA polymerase, RuvA, RuvB

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In DNA recombination repair DNA polymerase, RuvA and RuvB are associated with what step(s)?

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3. 3' end extension: branch migration
4. 3' end extension

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Enzymes?

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Define concatenation?

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replication of a circular chromosome results in two topologically interlinked (cantenated) circular chromosomes. DNA circles linked in this way are known as CATENANES.