Section 7: Molecular Genetics Flashcards
Describe the structure of DNA
Double helix with major and minor grooves
This carries the hereditary information of the cell
DNA
What is the DNA backbone made of?
5’ to 3’ phosphodiester bonds form a phosphate backbone
What are the nitrogen bases in DNA?
In RNA?
ATGC in DNA
AUGC in RNA
DNA replication begins at special sites in the middle of the DNA molecule (not the end) called
DNA strands separate to form __________ that expand in both directions
Thousands of them happen to speed up relication of 3 billion BP molecule in eukaryotic cells. How many origins of replication do prokaryotes have?
Origins of replication
Replication bubbles
One
In DNA replication, when is the second chromatid containing a copy of DNA assembled?
Interphase (S phase?)
During interphase, DNA is unzipped and each strand serves as a template for complementary replication
This is term means one strand of the two is old, the other is new, occurs in DNA replication of all known cells
Semi Conservative Replication
What is the enzyme that unwinds DNA?
It forms a Y shaped replication fork
Helicase
Single stranded binding proteins attach to each strand of uncoiled DNA to keep them separate
What break and join the double helix, allowing the prevention of knots?
(if you unwind a twist, the ends will get extra tight and knot up)
Topoisomerases
In what direction does DNA polymerase move?
Is the new strand parallel or antiparallel?
3’–>5’
Antiparallel (5’—->3’)
On this strand, the DNA polymerase works continuously as more DNA unzips
In this strand (the 5’–>3’ strand), the DNA polymerase has to go back to the replication fork and work away from it
What are the fragments that it produces called?
What connects these fragments?
Leading Strand
Lagging Strand
Okazaki Fragments
DNA ligase
In DNA replication, this is an enzyme that creates a small strip of RNA primer off which DNA polymerase can work since it can only add to an existing strand
Primase
What does DNA replication require at the start?
RNA primer
Every okazaki fragment has an RNA primer, these RNA strips are later replaced by…
DNA Polymerase 1
Other than replacing BPs from RNA primers, what else does DNA pol 1 do?
This polymerase is pure replication
DNA repair
DNA polymerase 3
DNA poly 1 and 3 have 3’–>5’ _____, it breaks phosphodiester backbone on a single strand of DNA and removes a nucleotide. It can only remove from 3’ end (in this case_ of the chain
Exonuclease
Which can also do proofreading, DNA poly 1 3, or both?
Which also has a 5’–>3’ exonuclease to take off the primer
both
Pol 1
in sum:
DNA Pol ______ mainly replicates the DNA 5’ to 3’ but can also proofread via 3’ to 5’ exonuclease.
DNA Pol ______ primarily breaks down RNA primer via 5’ to 3’ exonuclease and replaces it with DNA (laid down between Okazaki fragments mainly) via 5’ to 3’ polymerase while proofreading as it goes, can proofread
via 3’ to 5’ exonuclease as well
DNA polymerase 3
DNA polymerase 1
In prokaryotes, what happens to the good strand if there is an error in replication, so it doesn’t repair the wrong strand?
It is methylated
In all cases of repair, what must come seal the backbone afterward?
Ligase
Energy for elongation is provided by two additional ________ attached to each new nucleotide. Breaking the bonds
holding the two extra _________ provides chemical energy for the process (same w/ transcription!). Human rate 50n/s
Phosphates (both blanks)
A region of repetitive nucleotide sequences at each end of a chromatid, which protects the end of the chromosome from deterioration or from fusion with neighbouring chromosomes.
telomere
What two problems can occur in the replication of a telomere?
Not enough template strand where primase can attach
The last primase is removed
Do prokaryotes have telomeres?
No, circular DNA
enzyme that attaches to the end of template strand and extends the template strand by adding short sequence
of DNA over and over (not important code), allowing elongation of lagging strand to continue
For reference: https://www.youtube.com/watch?v=AJNoTmWsE0s
Telomerase
Telomerase carries an
RNA template: binds to flanking _____ end of telomere that compliments part of its RNA template, synthesizes to fill in over
the rest of its template
binds to 3’ end
For reference: http://upload.wikimedia.org/wikipedia/commons
/3/3b/Working_principle_of_telomerase.png
In protein synthesis, one-gene-one-polypeptide hypothesis defines a _____ as the DNA segment that codes for a particular polypeptide.
Also, genetic code is universal for nearly all organisms and most AAs have more than one codon specifying them. This concept is called
gene
redundancy/degeneracy
This type of RNA is a Single stranded template. Since there are 64 possible ways (4x4x4) ways that four nucleotides can be arranged
in triplet combinations, there are 64 possible codons. 3 of them are stop codons. Therefore, only 61 codes for amino acids
mRNA
This type of RNA has a C-C-A-3’ that attaches to an AA, and the other portion is the anticodon which base pairs with the codon in mRNA
tRNA
What part of tRNA base pairs with the codon of mRNA?
What fragment of tRNA attaches to an AA?
anticodon
C-C-A-3’
This is the term for the concept that the exact bp of the 3rd nucleotide in the anticodon and the 3rd nucleotide in the codon is often not required, allowing 45 different tRNA’s base-pair with 61 codons that code for amino acid. There is space for the 5’ anticodon on tRNA to move or…
Wobble
This type of RNA transports an AA to its mRNA codon
tRNA
Together w/ proteins, this type of RNA forms ribosomes
How many binding sites do ribosomes have? What are they?
rRNA
3, one for mRNA, tRNA that carries a growing polypeptide chain (Psite) and one for 2nd tRNA that delivers the next aa (A site)
The nucleolus is an assemblage of DNA being actively transcribed into..
rRNA
What are the termination sequences on mRNA?
UAA, UGA, UAG
Where is the ribosome assembled?
Where is it exported?
The nucleolus
Cytoplasm (large and small subunits separately)
The is the creation of RNA molecules from a DNA template
Prokaryotes are MONOCISTRONIC/POLYCISTRONIC
Eukaryotes are MONOCISTRONIC/POLYCISTRONIC
Transcription
Prokaryotes POLYCISTRONIC (each mRNA can create multiple polypeptide chains)
Eukaryotes MONOCISTRONIC (each mRNA creates a single polypeptide chain)
In this phase of transcription, the RNA polymerase attaches to a promoter region on DNA and unzips the DNA into two strands
The promoter region often contains what sequence?
Initiation
TATA (TATA box)
The most common sequence of nucleotides at the promoter region is called the _____ sequence
Variations from it cause what?
Consensus
less tight RNA polymerase binding and a lower transcription rate
In this phase of transcription, RNA polymerase unzips DNA and assembles RNA nucleotides using one strand of DNA as template.
Elongation
only ONE strand is transcribed!
This phase of transcription occurs when RNA pol reaches a special sequence, often AAAAAAA in eukaryotes
Termination
In what direction of the DNA strand is transcription occurring?
3’ to 5’ (synthesized RNA is 5’ to 3’)
In mRNA processing, before leaving the nucleus, pre-mRNA undergoes several modifications
First this sequence is added to the 5’ end of the mRNA. It’s a guanine with three phosphate groups, also called ___, providing stability at the point of attachment for ribosomes
5’ cap (-P-P-P-G-5’)
GTP
In mRNA processing, before leaving the nucleus, pre-mRNA undergoes several modifications
After the 5’ cap is added, this sequence is attached to the 3’ end of the mRNA. It provides stability and control of movement across the nuclear envelope
How many AAs is it?
A poly-A tail (AAA…AAA-3’)
200
What does the A poly A tail facilitate in prokaryotes?
Degradation
After the A poly A tail is attached to the 3’ end of the mRNA, this occurs to remove nucleotide segments from mRNA, deleting introns and splicing exons
RNA splicing
During DNA splicing in mRNA processing, before the mRNA moves into the cytoplasm, what delete the introns and splice the exons?
Do prokaryotes have introns?
Small nuclear ribonucleoproteins (snRNPs) and splicosome
No
After RNA splicing in mRNA processing, this occurs, which allows different mRNA to be generated from the same RNA transcript by selectively removing differences of an RNA transcript into different combination, coding for a different product
Alternative Splicing
EUKARYOTES/PROKARYOTES generally have ready to go mRNA upon transcription
Only in EUKARYOTES/PROKARYOTES do you need mRNA processing
Prokaryotes have ready to go mRNA, translation can occur immediately
eukaryotes require proessing,
In which can multiple RNA polyermases transcribe the same template simultaneously, prokaryotes, eukaryotes, or both?
Both
This process is the assembly of AAs based on reading of new RNA
What is the energy source?
Translation
GTP
This is located in the cytoplasm for translation, the amino acid attaches to it at the 3’ end requiring 1 ATP
Aminoacyl-tRNA
In this phase of translation, the small ribosome unit attaches to the 5’ end of mRNA, the tRNA-methionine attaches to the start sequence of mRNA and the large ribosomal unit attaches to form a complete complex
What is the start sequence of mRNA
Initiation
AUG
During the initiation of translation, the small ribosomal subunit attaches to what end of mRNA?
The large ribosomal subunit binds…
For reference: http://highered.mcgraw-hill.com/sites/9834092339/student_view0/chapter
15/translation_initiation.html
5’
The small subunit
During this step of translation, the 2nd tRNA binds to the A site for peptide bond formation. This 1st tRNA is released without methionine. The 2nd tRNA moves to the p site and the next tRNA comes into A site to repeat process
Elongation
The movement of tRNA from the A site to the P site during elongation during translation is referred to as
Translocation
During this step of translation, the complex encounters a stop codon (UAG, UAA, UGA). The polypeptide and two ribosomal units all release once the release factor breaks down the bond between tRNA and the final AA of the polypeptide.
Termination
While a polypeptide is being translated, AA sequences determine the folding conformation. Folding is assisted by
Chaperone Proteins
After translation, signal peptide at the beginning of the translated polypeptide may
direct the ribosome to attach to the ER, in which case the polypeptide is injected into the
If injected, the polypeptide may be secreted from the cell via
ER lumen
Golgi
In general, post translational modifications may occur with the addition of sugars, lipids, phosphate groups, etc. They may be subsequently processed by the ___ before it is functional
Golgi
Amino Acids are placed starting from the ____ end of the mRNA and move all the way down to the ____ end
tRNA codons for matching are ___ to _____
5’ end, move all the way down to the 3’ end
tRNA codons for matching 3’ to 5’
Multiple ________ may simultaneously translate 1 mRNA
Ribosomes
In bacteria the start codon is _____ rather than methionine
n-formylmethionine
This is a mutation where the new coon still codes for the same AA
This is a mutation where there is no change in protein fxn
Silent mutation
Neutral mutation
his is a mutation where the new codon codes for a stop codon
Nonsense mutation
This is a mutation where a new codon codes for a new AA. Can be minor or fatal, like sickle cell (val substituted for glu)
Missense mutation
What are the three repair mechanisms?
Proofreading
Mismatch repair
Excision repair
Proofreading of DNA is conducted by _______ which checks base pairs
This type of DNA repair has enzymes repair things DNA polymerase missed
This type of repair has enzymes remove nucleotides damaged by mutagens
DNA polymerase
Mismatch repair
Excision Repair
This is DNA coiled around bundles of 8/9 histones like beads on a string
Nucleosome
When it’s not division, chromatin exists as either of two types
This type is loosely bound to nucleosomes, actively being transcribed
This type is areas of tightly packed nucleosomes where DNA is inactive
Euchromatin
Heterochromatin
Heterochromatin has a lot of ____________, which are large tandem repeats of noncoding DNA
Satellite DNA
There are DNA segments that can move to new location on same/different chromosome
Transposons
This type of transposon consist of only one gene that codes for enzyme that just transports it (transposase)
These code for extra: replication, antibiotic resistance, etc.
Insertion of transposons into another region could cause…
Insertion Sequence
Complex transposons
mutation (little to no effect)
What are the 3 parts of a virus
Nucleic acid
Capsid
Envelope
In viruses, can the nucleic acid be DNA, RNA or either?
Can it be double stranded, single stranded, or either?
It can be one or the other
It can be one or the other
(remember big table in microbio?)
This is the protein coat that encloses the nucleic acid
What assemble and form the nucleic acid?
Capsid
Capsomeres
This part of the virus surrounds the capsid of some viruses, it encorporates phospholipid/protein obtained from cell membrane of host
Envelope
This is a virus that only attacks bacteria
Bacteriophage
Are viruses usually specific or nonspecific to type of cell and species?
Highly specific
This is the range of organisms that a virus can attack
Host range
In this type of viral replication, this occurs when the virus penetrates the cell membrane of the host and uses host machinery to produce nucleic acids and viral proteins that are then assembled to make new viruses
The viruses burst out of the cell and infect other cells
Lytic Cycle
In a DNA virus, DNA is replicated and forms new viral DNA which is transcribed to produce…
Viral proteins (they assemble to form new viruses)
For this type of virus, RNA serves as mRNA, it is translated into protein
RNA virus
These are viruses that use reverse transcriptase to make a DNA complement of their RNA, which can manufacture RNA or enter the lysogenic cycle
Retroviruses
In this type of viral replication, viral DNA is incorporated into the DNA of the host cell
When in this state, it remains inactive until external stimuli triggers it to enter the lytic cycle
What the virus called if it enters lysogenic cycle?
Lysogenic Cycle
Dormant State
Provirus or prophage (bacteria)
What are on the cell wall of bacterium that are used as recognition and binding sites by bacterial viruses? They also provide cell wall rigidity
What type of bacteria are they found on?
Techoic Acids
ONLY on gram positive
These are not viruses or cells. They are misfolded versions of proteins in brain that cause the normal proteins to misfold as well. They are fatal.
Prions
How do bacteria reproduce?
Binary fission
This type of replication has the chromosome replicating and the cell dividing into two cells, each with one chromosome
Do bacteria have a nucleus? Microtububles? Spindle? Centriole?
Binary Fission
Nope not needed for replication
These are short, circular DNA segments outside the chromosome. They carry genes that are beneficial but not necessary for survival
Plasmids
Do plasmids replicate with the host cell?
These are plasmid that can integrate into bacterial chromosome
No, they replicate independently
Episomes
This is a method of genetic variation among bacteria where one bacterial cell produces a bridge (pilus) and connects to a recipient bacteria. It sends chromosome/plasmid and recombination to occur
Conjugation
The conjugative plasmid, it is an episome that can integrate itself into the bacterial chromosome..
Once the recipient receives it, it is now ____ and can donate as well
This type of plasmid provide bacteria with antibiotic resistance
F Plasmid/F factor
F+
R plasmids
What can be used for cell adhesion in bacterial cells?
Pili
This method that produces genetic variation in bacteria introduces DNA into the genome by a virus via the lytic cycle.
When the virus infects another host, the bacterial DNA part that it delivers can recombine with resident DNA
Transduction
This method of genetic variation in bacteria where bacteria absorb DNA from surroundings and incorporate it into its genome
Transformation
In the regulation of prokaryotic gene expression, these control gene transcription
What do they consist of? 4 things
Operons
Promoter
Operator
Structural Genes
Regulatory Genes
This part of the operon (prokaryotic gene expression regulation) is a sequence of DNA where RNA polymerase attaches to begin transcription
Promoter
This part of the operon (in prokaryotic regulation of gene expression) is the region that can block action of RNA polymerase if occupied by a repressor protein
Operator
This part of the operon (prokaryotic regulation of gene expression) is DNA sequences that code for related enzymes
This part is located outside of the operon region, produces repressor produces and activator proteins which assist in the attachment of RNA polymerase to promoter region
Structural Genes
Regulatory Genes
In the regulation of prokaryotic gene expression, this is an operon that has a regulatory gene that controls the breakdown of lactose
The regulatory gene produce an active repessor called ______ and blocks RNA polymerase. When lactose is available, it binds the repressor and inactivates it, RNA polymerase can now transcribe.
So the operon is induced by
The enzymes that the operon produces are said to be…
Lac Operon (E. coli)
Bind operator
Lactose
Inducible enzymes
Lac operon consists of three lac genes, Z, Y and A which code for…..
B-galactosidase, lactose perease, and thiogalactoside thransectylase
In lac operons, low gluose means high what levels? It binds to CAP binding site of promoter so RNA polymerase more efficiently transcribes in high lactose levels
cAMP
This operon produces an enzyme for trp synthesis
Regulatory genes prouce and inactive repressor, RNA polymerase produces enzymes. When trp is available, you no longer need to synthesize it internally. It binds to the inactive repressor and inactivate it. It then binds the operator and blocks RNA polymerase
Trp is a _______
Trp operon
corepressor
When structural genes stop producing enzymes only in presence of active repressor, the enzymes are called
Repressible enzymes
Unlike some repressible enzymes, some genes are _____ (constantly expressed) either naturally or due to mutation
Constitutive
In the regulation of eukaryotic gene expression, these are repressors and activators, they influence RNA polymerase’s attachment to the ____ region
Regulatory Proteins’
Promoter region
In the nucleosome packing in regulation of eukaryotic gene expression, this induces tighter packing prevent transcription
This uncoils the nuclesome and allows transcription to proceed
Methylation of histones (also used in X inactivation on DNA bases to repress DNA activity)
Acetylation of histones
These block mRNA transcriptions, translation, or degrade existing mRNA in eukaryotic gene expression
This is mRNA which folds back into itself. It gets chopped up then made single stranded. The relevant strand will bind to DNA (prevent transcription) or mRNA (signals destruction)
Short interfering RNAs (siRNA)
dsRNA (double stranded RNA)
What percent of the human genome does not code for a protein product?
97%
Noncoding DNA includes
regulatory sequences, introns,
repetitive sequences never transcribed, etc. Tandem repeats abnormally long stretches of back to back repetitive sequences
within an affected gene (e.g. Huntington’s).
Tandem repeats are abnormally large back to back repetitive sequences within an affected gene, and play a role in this disease when expanded past normal.
Huntington’s Disease
This DNA contains DNA segments or genes from different sources. It can come from viral transduction, bacterial conjugation, transposons, or artificial technology
Recombinant DNA
Crossing over during the ____ phase of meiosis produces recombinant moleculesPro
Prophase
For recombinant DNA, technology uses ________ to cut up specific segments of DNA and leave it with a ____ end (unpaired)
Restriction endonucleases
Sticky End
These restriction enzymes are normally used by bacteria to protect against viral DNA
They protect their own DNA via…
EcoRI, BamHI
methylation
This is a vehicle used to transfer foreign genetic material to another cell
What is an example of a DNA molecule that can be a vector?
Vector
Plasmid
To introduce foreign DNA into a plasmid, the plasmid is treated with the same ________ so the same stick ends bind from the previously cleaved DNA molecule
What stabilizes the attachments?
Then the plasmid is introduced into bacterium by
Restriction enzyme
DNA ligase
Transformation
In order for bacterium to take up a modified plasmid (treated with restriction enzymes), it must be made competent with
After this process, bacteria can be grown to produce product. What can be used to filter out the ones that don’t have recombinant DNA?
Electroporation or heat shock + CaCl2
Antibiotic resistance/screen method
This is a technique that uses agarose gel under an electric field for the separation of proteins based upon charge and size
DNA moves toward the anode or cathode?
Does shorter DNA move further or less than longer DNA?
Gel electrophoresis
DNA moves towards the positively charged anode (DNA is a negative molecule, ANIONS MOVE TOWARDS THE ANODE)
Further
Can electrophoresis be done on proteins?
Yes. they have negative charges and positive charges to separate them based upon charge.
These can be identified between individuals, it is often used in paternity suits or to identify crime suspects as it’s a result of Mendelian genetics
Restriction Fragment Length Polymorphism
restriction fragments are compared and differ in length because of polymorphism
This is used at crime scenes to compare the RLPs of suspects
DNA fingerprinting
This is a repeat of 2-5 nucleotides that differ between all individuals except identical twins
STR (short tandem repeat)
this enzyme makes DNA molecule directly from mRNA
DNA obtained in this manner is complementary DNA which lacks ______ that supresses transcription
Reverse Transcriptase
Introns
This technique uses synthetic primer (the primer may be RNA or DNA oligonucleotides) to clone DNA (rapidly amplify). Taq polymerase (heat stable) + nucleotides + primers + salts (buffer) necessary.
PCR
What are the 3 steps of PCR
- Denaturation (>90degreesC)
- Add primers + Anneal
- Elongation (taq polymerase)
This is a technique used to ID target fragments on known DNA sequences in a large population of DNA. Electrophoresis is first, then the DNA fragments are transferred to nitrocellulose membrane, then a probe is added which will hybridize and mark it
Southern Blotting
This is a technique that is the same as Southern blotting but is used for RNA fragments
Northern blotting
This is similar to Southern blotting but for proteins. Electrophoresis, blot to membrane, primary antibody specific to protein added to bind to
that protein, then secondary anti-body-enzyme conjugate will bind to primary and mark it w/ enzyme for visualization
Western Blotting
Note: an easy way to identify all of the blotting methods is to use the phrase SNOW DROP. Match the letters in their order for each word, skipping O in both words
SNOW
DROP
The genome of humans differs roughly one of every _____ nucleotides
These differences are called
Nucleotide polymorphisms (SNPs)
Plasmids are circular DOUBLE/SINGLE stranded DNA
They have ______ sites for restriction enzymes.
If these sites are cut, what is the shape of the DNA?
Double stranded DNA
Restriction sites
linear
Do plasmids get reproduced during cell division?
It depends! They can or they cannot.
Do bacteria readily take up plasmids?
No, that’s why there’s antibiotic resistance screening to clone the one’s that actually did take it up.
If you’re making a prokaryotic gene product in a mammalian cell, what do you need to add for the mRNA to survive
If making a eukaryotic gene product in a prokaryotic cell, what do you need to make sure to not have?
PolyA tail (look it up)
Introns
This is the term for complementary bases annealing
Hybridization
If you want to test for a specific gene sequence in someone’s DNA, this is the first method in the notes, testing for a single gene only.
take drop of blood, cut up DNA, use PCR
method w/ specific primer for that region. If that gene is there lots of copies. Gene not there —> no copies.
This is a second method of testing for a specific gene in someone’es DNA, and can test for many things at the same time.
take drop of blood, PCR it to amplify. We have a solid support w/ pieces of ssDNA w/ specific sequence covalently attached –> will hybridize to anything complementary (e.g. disease genes). On that same solid support we can put sSDNA pieces specific for other genes, can do this hundreds of times at different spots on this DNA microarray. Take desired amplified DNA, heat it to denature, add to DNA microarray, any DNA that hybridizes is a match. The amplified DNA we added is already fluorescently tagged –> hybridization wash to get rid of weakly bound sequences (e.g. not a complete match) –>add a dye that will show heavily if something has bound to our microarray. Can also do this starting w/ mRNA –>reverse transcriptase –>PCR amplify –>etc.
What can cleave the phosphodiester backbone to chunk out NTs in DNA?
What cut out just the NTs?
endonucleases
Exonucleases
This term means that a cell can give rise to any and all human cells, and even an entire functional organism
This term means a cell can give rise to a limited range of cells within a tissue type
Totipotent
Multipotent
