Term
| Why is the human genome much larger than bacterial? |
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Definition
| Bacteria genes use less DNA sequences meaning they contain less base pairs than an analogous gene |
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Term
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Definition
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Term
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Definition
| Cytosine, uracil, thymine |
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Term
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Definition
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Term
| B-DNA has major and minor groove and DNA-binding proteins bind in major groove |
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Definition
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Term
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Definition
| number of helical turns of one strand around the other, coil |
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Term
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Definition
| number of times double helix crosses over itself (supercoiling) |
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Term
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Definition
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Term
| DNA is a right-handed helix |
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Definition
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Term
| Why is most DNA negatively supercoiled? |
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Definition
| Causes underwinding which makes it easier to separate the strands for replication |
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Term
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Definition
• Type I nicks only one strand of the DNA
• Type II directly alters number of writhes by passing dsDNA segment through another |
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Term
| Why do type II topoisomerases require ATP? |
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Definition
| These topoisomerases have to undergo a conformation change in order to cleave a DNA molecule and hold the two strands apart. Energy needed from ATP is required to have the enzyme revert back to its original conformation. |
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Term
| Importance of topoisomerases in replication |
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Definition
| Essentially they help unwind DNA (relieve supercoils) so that replication is possible |
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Term
| General mechanism for Topoisomerase I |
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Definition
Human topo has four domains that encircle the DNA. The surface of the protein that contacts the DNA is rich in positively charged groups that can interact with the backbone phosphate groups of about 10 base pairs. When one strand of DNA is cleaved, an active-site Tyr residue forms a covalent bond with the backbone phosphate at one side of the nick.
Remains bound to DNA as passes strand through |
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Term
| What does semi-conservative mean? |
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Definition
| This means that for every round of replication, there will be two copies of DNA, with one copy a parental strand, and the other copy a daughter strand. |
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Term
| General model of replication |
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Definition
| Factory model: Machinery stationary and DNA feeds thru |
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Term
| Proofreading ability of DNA pol |
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Definition
| • DNA pols contains a second active site that catalyzes hydrolysis of the wrong nucleotide at the 3' end of the growing DNA strand. So in essence, it has 3'->5' exonuclease activity |
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Term
| Where does energy come from to form phosphodiester bond when DNA pol adds nucleotides? |
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Definition
| • It comes from hydrolysis of the high-energy phosphate bonds between the phosphates attached to the base to be added |
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Term
| General structure of DNA pol |
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Definition
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Term
| Significance of MG2+ ions in active site of DNA pol? |
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Definition
• Helps to neautralize the negative charges on the phosphodiester backbone
• 1 Mg2+ ion interacts with 3' O atom to make it more nucleophilic so it's more likely to attack the phosphate of the incoming nucleotide |
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Term
| Why won't rNTPs bind to DNA pol? |
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Definition
| • The deoxy carbon at the 2' position of the nucleotide lies in a hydrophobic pocket. This binding site allows the polymerase to discriminate against ribonucleotides which bear a 2' OH group |
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Term
| Is there more than one DNA polymerase? Why? |
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Definition
| • Yes because some of them perform specialized roles and in eukaryotes, two different types work in concert to synthesize the leading and lagging strand |
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Term
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Definition
• Relax supercoiling by nicking one strand (Topoisomerase I)
• Helicase – Unwinds helix (separates 2 strands)
• Need proteins to protect ssDNA from nuclease digestion
o SSB: single-strand binding proteins
• RNA primers synthesized
• DNA replicated
• RNA removed and replaced with DNA
• Gaps sealed
• Supercoiling added |
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Term
| The difference between the leading and lagging strand – why is there a difference and how is this dealt with in the process of replication? |
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Definition
| • Leading strand is synthesized continuously while the lagging strand is synthesized in pieces using multiple RNA primers and it contains Okazaki fragments. There is a difference because the two DNA strands are antiparallel. This is dealt with by having two polymerases work side-by-side while one template DNA strand is periodically looped out. |
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Term
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Definition
| o coats the single stranded DNA to protect it from nucleases and to prevent them from reannealing or forming secondary structures that might impede replication. |
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Term
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Definition
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Term
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Definition
| o Is a 5'-3' exonuclease that removes RNA primer |
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Term
| General mechanism of DNA ligase |
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Definition
| • The discontinuous segments of the lagging strand are joined by the action of DNA ligase. It works by activating the 5' phosphate on DNA with AMP which is transferred from ATP in eukaryotes and NAD in prokaryotes. |
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Term
| Why are telomers necessary? |
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Definition
| They are necessary to counteract the potential loss of genetic information |
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Term
| How are telomers synthesized? |
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Definition
| Telomerase extends the 3' end of a DNA strand by adding TTAGGG repeats. DNA polymerase can then extend the complementary strand by the normal mechanism for lagging-strand synthesis. |
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Term
| General mechanism of telomerase |
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Definition
| • Uses an enzyme-associated RNA molecule as a template, is a reverse transcriptase. |
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Term
| How the ssDNA regions of the telomere are protected from nuclease digestion during replication |
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Definition
| • Telomeric DNA folds back on itself to protect it |
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Term
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Definition
| • Occurs when the base pairs are not mismatched, not damaged. |
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Term
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Definition
| • Cellular metabolism itself exposes DNA to the damaging effects of reactive oxygen species. When the modified DNA strand is replicated, it can lead to nucleotide substitutions |
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Term
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Definition
| • Glycosidic bond connecting base to sugar is broken |
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Term
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Definition
| • amine group is removed which can alter the identities of bases |
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Term
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Definition
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Term
| During mismatch repair, how does the repair machinery know which strand is wrong? |
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Definition
| • Bacterial MutS protein binds to the mismatched pair and causes the DNA to bend. This induces an endonuclease to cleave the strand with the wrong nucleotide. Endonuclease recgonizes which strand is new by determining if it's methylated or not. Methylation occurs after replication and marks the strand as being mature. |
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Term
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Definition
• Occurs when the actual bases themselves are damaged.
• DNA glycosylase binds to DNA and removed damaged base
• AP endonuclease nicks the backbone
• DNA pol fills in gap, ligase seals nick |
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Term
| Nucleotide excision repair |
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Definition
| • UV damage results in thymine dimers. Mechanism is similar to excision repair except the whole segment containing dimer + 30 nt is cut off. Filled in by DNA pol; nick sealed by ligase |
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Term
| Nonhomologous end joining |
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Definition
• Broken DNA recognized by Ku protein
• Ku undergoes conformation change and recruits nuclease
• Nuclease trims residues from end
• Polymerase recruited that utilizes template-independent polymerization |
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Term
| Why is nonhomologous end joining most common even though it's inherently mutagenic? |
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Definition
| • It's the lesser of two evils vs damaged DNA that can be passed on |
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Term
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Definition
o Intact strand serves as template for broken strand
o Must have homologous DNA |
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Term
| How is cancer a genetic disease? |
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Definition
| • It’s a genetic disease because all the known causes of cancer are linked in some way to damage of the cell’s DNA so cancer is fundamentally a disease of the genes. |
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Term
| What are the two general ways that cancer can be stimulated? |
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Definition
• Oncogenic mutations that cause cells to grow and divide even in the absence of a growth signal.
• Inactivating genetic events such as the inactivation of a tumor suppressor gene. |
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Term
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Definition
| • P53 stimulates production of a protein that blocks the cell’s progress toward cell divison which gives the cell time to repair DNA |
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Term
| How does p53 level control response of cell and balance between repair and cell death |
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Definition
| o The level of p53 in the cell is controlled by its rate of degradation. A little bit of p53 in the cell will lead to repair but if there’s a lot of p53, this signals that the cell is damaged beyond repair and so cell death will be appropriate. |
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Term
| Why is p53 often associated with tumor growth? |
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Definition
| o Because without it, cells can’t repair themselves or kill themselves. |
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Term
| difference between heterochromatin and euchromatin |
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Definition
• Heterochromatin is densely packed, stains intensely. Because it’s so densely packed, it’s not suitable for transcription.
• Euchromatin is less condensed, doesn’t stain as well and is more suitable for transcription. |
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Term
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Definition
| • It consists of 8 histone proteins (4 pairs) with DNA wound around it. |
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Term
| How is the nucleosome further packaged? |
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Definition
| it is further packaged into the 30-nm fiber and even more higher levels of packaging |
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Term
| How histones are modified? |
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Definition
| • Acetylation promotes transcription because it weakens histone histone interaction with DNA while deactylation does the opposite. |
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Term
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Definition
| • Addition of a methyl group can alter interactions with DNA-binding proteins |
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Term
| Significance of CpG islands |
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Definition
| • Cpg islands are regions with a high frequency of CpG sites, and they are usually located near the starting point of genes. If these sites get methylated, this can “silence” that particular DNA segment. |
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Term
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Definition
o DNA is first denatured to produce single stranded DNA
o A primer, DNA pol, dNTPs, and labeled ddNTPs are added
o DNA chains of various lengths are created and these chains are sent through gel electrophoresis to determine the sequence of the temple strand. |
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Term
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Definition
o Is a reaction that amiplifies DNA.
o Has 3 basic steps:
• Strand separation
• Primer binding
• Primer extension
• Basically, DNA is denatured, a primer is attached to both strands and the complementary strands are denatured again to repeat the process |
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Term
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Definition
| o Cuts DNA at specific sequences where palindromes are located |
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