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83 Cards in this Set
- Front
- Back
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How was DNA determined as the genetic material? |
heat killing different virulent strains, the Hershey Chase experiment with T4 phage |
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What are the 3 key properties of DNA? |
must allow faithful replication, must have informational content, must be able to change |
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What are the building blocks of DNA? |
phosphate, deoxyribose sugar, nitrogenous base |
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What are purines? |
adenine and guanine, 2 ring structure |
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What are pyrimidies? |
cytosine and thymine, one ring structure |
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What are the chemical components of DNA arranged into? |
nucleotides |
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What is Chargaff's rule? |
A-T, C-G, T+C=A+G |
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What holds the double helix together? |
hydrogen bonds between the bases of the strands |
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What forms the backbone of DNA? |
alternating phosphate and deoxyribose units connected by phosphodiester linkages |
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Where do phosphodiester bonds form? |
5' carbon of one deoxyribose to the 3' carbon of the adjacent deoxyribose |
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Where are the bases attached? |
the 1' carbon of the deoxyribose sugar |
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What is the most stable form of base stacking? |
double helix with two grooves running in a spiral with a major groove and a minor groove |
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Where is most DNA-protein associations? |
the major groove |
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Which way does DNA move? |
right handed |
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How many H bonds on base pairing? |
3 in G-C, 2 in A-T |
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What do the strands act as? |
each strand acts as a template for complementary base pairing |
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What is the semiconservative mechanism of replication? |
each daughter molecule should have one parent nucleotide chain and one newly synthesized one |
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What is conservative replication? |
the parent DNA molecule is conserved and a single daughter double helix is produced |
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What is dispersive replication? |
daughter molecules have strands each with parts of both parental and newly synthesized DNA |
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What is the Meselson-Stahl experiment and what did it show? |
way of discovering the replication mechanism through cesium chloride gradient centrifugation where the nitrogen of DNA had different isotopes and the DNA was separated by isotope in E Coli |
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What is the replication fork? |
the location at which the double helix is unwound to produce the two strands for templates |
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What does DNA polymerase do in DNA? |
adds deoxyribonucleotides to the 3' end of the growing nucleotide chain |
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What are the substrates for DNA polymerase? |
dATP, dGTP, dCTP, dTTP |
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What is the addition of each base accompanied by? |
removal of 2 of the 3 phosphates in the form of pyrophosphate (PPi) |
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How many polymerases are in E Coli? |
5 |
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What does DNA polymerase I, or pol I, do? |
catalyzes chain growth in the 5-3' direction, acts as a 3-5' exonuclease activity which removes mismatched bases, and a 5-3' exonuclease activity which degrades single strands of DNA or RNA |
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Why was pol I not expected to play a big part in DNA synthesis? |
it was too slow (20 nuc/second) and too abundant (400 molecules/cell) |
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What DNA polymerase catalyzes DNA at the replication fork? |
pol III |
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Where does DNA pol III act? |
the replication fork |
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Why can only one strand serve as a template in the direction of the replication fork? |
DNA polymerase can only add to the 3' end so synthesis can only occur this way on one strand because DNA runs antiparallel |
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What is the leading strand? |
newly synthesized strand that forms in the direction of the replication fork |
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What happens in the other strand? |
the second segment is synthesized in short stretches called Okazaki fragments |
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What must initiate synthesis of both strands? |
a primer which is a sort chain of nucleotides that bind with the template strand to form a segment of duplex nucleic acids which helps DNA start the chain |
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What are primers synthesized by? |
a set of proteins called a primosome with an enzyme called primase |
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What does primase do? |
synthesizes a short strand of complementary RNA to a specific stretch of the chromosome |
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How many primers are needed on the leading strand? |
1 |
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How many are needed for the second strand? |
one primer for each Okazaki fragment |
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What does pol I do during DNA replication? |
removes the RNA primers and fills in the gaps |
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What does DNA ligase do? |
joins the 3' end of the gap filling DNA to the 5' end of the downstream Okazakif ragment |
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What is the new strand called? |
the lagging strand |
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How does DNA ligase work? |
joins broken pieces of DNA by catalyzing the formation of phosphodiester bonds between adjacent fragments |
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What is the error rate of DNA replication? |
called fidelity or accuracy. less than one error per 10^10 nucleotides |
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What helps with DNA replication accuracy? |
pol I and III have a proofreading function and can get rid of mismatched bases |
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What often causes the addition of an incorrect base? |
tautomerization where different isomers are placed --> the keto form is the normal form but the bases can shift to the imino or enol form which can form a mispair |
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Which is more likely to have errors, RNA or DNA? |
the RNA primer is more likely to have errors because primase does not have proofreading activity |
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What is the repliosome? |
the nucleoprotein complex that coordinates the activites at the replication fork |
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What is the pol III holoenzyme? |
two catalytic cores and accessory proteins which handle synthesis of each strand with accessory proteins that bridge the two catalytic cores |
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What is the beta clamp? |
an important accessory protein that encircles the DNA like a donut and keeps pol III attached to the DNA molecule to allow continuous synthesis |
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What is a distributive enzyme? |
an enzyme that can only add 10 nucleotides before falling off |
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What is a processive enzyme? |
an enzyme that can stay at the moving fork and add tens of thousands of nucleotides |
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What does primase act as? |
a distributive enzyme |
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What opens the double helix? |
helicases |
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What prevents overwinding? |
topoisomerases |
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What do helicases do? |
enzymes that disrupt hydrogen bonds that holds the double helix together by forming a donut around the DNA and unzips it |
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What do SSB proteins do? |
stabilize the unwound DNA by binding to the single strand and preventing the duplex from forming |
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What do topoisomerases do? |
relax supercoiled DNA by breaking either a single DNA strand or both strands which allows the DNA to rotate in a relaxed molecule and then rejoin the strands |
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Where does assembly of the replisome occur? |
on the origins of the chromosome |
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What is the first step in replisome formation? |
DnaA protein binds to a 12 base pair sequense (DnaA box) which is repeated five times in oriC (origin) |
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What is the second step? |
the origin is unwound at a cluster of A + T nucleotides |
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What happens after unwinding begins? |
additional DnaA proteins bind to the unwound single stranded regions where the two helicases can bind and begin unzipping the helix at the replication fork |
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What prompts pol III and primase? |
protein protein interactions |
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Is DnaA considered part of the replisome machine? |
no- it only brings the replisome to the right place |
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How many growing forks are in yeast and humans? |
yeast- 400, humans- thousands |
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What results at the end of replication? |
two identical daughter molecules of DNA |
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When does DNA synthesis occur? |
the S phase of the eukaryotic cell cycle |
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What happens in yeast replication? |
three proteins being assembly of the replisome, starting with the origin recognition complex that first binds to sequences in yeast which recruits Cdc6 and Cdc1, which recruits the MCM helicase coplex |
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How can the replisome be assembled before S phase? |
the proteins are synthesized during late mitosis and G1 so they may already be there before S phase and are then degraded by proteolysis after synthesis begins |
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When do gene regions replicate? |
gene rich regions during early S phase and gene poor regions during later S phase |
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How do ORCs work in humans? |
they have a higher affinity for origins in open chromatin and bind there first and then only later bind to the gene poor regions after the others have been replicated |
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What are the telomeres? |
the two ends of linear DNA molecules |
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What happens to the telomeres in lagging strand synthesis? |
requires primers in advance so sequences are missing at the end of the strand so a single stranded tip remains |
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What happens if the daughter chromosome with the single stranded tip replicates? |
it would then become a shortened double stranded molecule after replication |
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How do cells avoid telomere shortening? |
cells add multiple copies of a simple noncoding sequence to the DNA at the chromosome tips which shortens the repeating sequences and not the necessary parts and are then added back to the chromosome ends |
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What does telomerase do? |
adds the short repeats to the 3' end of DNA molecules with the help of an RNA molecule which acts as a template |
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What is the RNA sequence template? |
3-AAUCCC-5 |
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How does the RNA sequence addition work? |
the telomerase RNA anneals to the 3' DNA, which is then extended with the RNA template and the protein. then, the telomerase moves down to extend the chain |
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What is the notable feature in telomere replication? |
the RNA serves as a template for DNA synthesis --> called reverse transcriptase activity |
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What is telomerases' other function? |
preserves chromosomal integrity by forming protective caps which makes sure the ends are capped and not mistaken for double stranded breaks |
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Why are double stranded breaks dangerous? |
can result in chromosomal instability which can lead to cancer and aging issues |
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How are double stranded breaks dealt with? |
they are either fused to another break (senescence) or initiated into a cell death pathway (apoptosis) |
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Which cells produce little telomerase? |
somatic cells, but germ cells have ample telomerase |
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What is Werner syndrome? |
premature aging possibly caused by shorter telomeres due to a mutation in the WRN gene that encodes for the protein that forms the telomere cap |
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What happens with cancer cells? |
they have telomerase activity which may be why cancer cells can grow in cell cultures for long years but normal cells will die --> try to develop drugs that inhibit telomerase activity to prevent cell growth |
