Lecture 6 Flashcards
In triploid organisms, how do they become triploid (sterile)?
Polar bodies in meiosis II aren’t extruded
Most common non-LTR retrotransposons (retroelements)
LINE-1 and Alu
Eukaryotic cells were broken open at low or moderate salt concentrations
and the DNA was seen in this form, allowing what to be seen?
Nucleosomes
In real life, there is more ___ than protein
DNA
In regular chromatin, what percent is DNA?
50%
The most common nucleosome width is?
30 nm
At ___ salt concentrations, the
nucleosomes display a classic
beads-on-a-string morphology
low
Two proteins responsible for loops of the 30 nm fibers
SMC and CTCF
Where is heterochromatin located?
Half of it at the nuclear periphery and half of it internal
What happens is euchromatin is overexpressed?
Heterochromatin is lost quickly
A temporary copy of a gene that contains information to make a polypeptide
mRNA
Produces a polypeptide using the information in mRNA
translation
becomes part of a functional protein that contributes to an organism’s traits
polypeptide
What is the central dogma?
DNA replication (using chromosomal DNA)–> transcription (using mRNA)–> translation into polypeptide
Rosalind Franklin revealed what? Using what?
DNA as a helix shape using X ray diffraction
Watson and crick realized Rosalind Franklins discovery as DNA as a helix, was….
too wide to be single stranded
Who discovered the three ways DNA may replicate
Mathew Meselson and Frank Stahl
Both parental strands stay together after DNA replication
Conservative model
The double-stranded DNA contains one parental and
one daughter strand following replication
Semi-conservative model
Parental and daughter DNA are interspersed in both strands
following replication
Dispersive model
Watson and crick confirmed which way to replicate?
Semi-conservative
What experiment was done to distinguish between the three
models of DNA replication?
- Grew e. coli in the presence of 15N (heavy isotope of Nitrogen) for many generations.
- Switch to e coli medium containing only 14N
- Collect sample of cells
- Analyze the DNA density by centrifugation using a CsCl gradient
When analyzing the DNA from distinguishing the models of DNA replication, what DNA disappeared and was replaced with something ___. This proved what type of DNA replication.
heavier, lighter, semi-conservative
During the DNA analysis of distinguishing the DNA models of replication–After ~ two generations, DNA is of two types: ____. After one generation DNA is ____
light and half heavy (semi-conservative). half heavy (consistent with dispersive and semi-conservative)
In DNA replication, the two dna strands separate and each serves as a ___
template strand for the synthesis of new strands
In DNA replication, The two newly made strands =
daughter strands
When a DNA strand is synthesized, nucleotides are only added onto the ___ end of the strand
3 prime
Bacterial DNA Replication
as seen by the
electron microscope
In bacterial DNA, there is one origin or replication and the polymerases meet where?
At opposite ends
The origin of replication in E. coli is termed
oriC (origin of Chromosomal replication)
Synthesis of DNA proceeds ___ around the bacterial chromosome
bidirectionally
The replication forks eventually meet at the ____ side of the bacterial chromosome
opposite
E. Coli OriC and the replication origins of other bacteria have ___ DNA sequences
highly conserved
DNA replication is initiated by the binding of ___ to the DnaA box sequences
DnaA proteins, DNaA box sequences
The binding of DnaA proteins to the DnaA box sequences, stimulates what?
binding of an additional 20 to 40 DnaA proteins to form a large complex
What other proteins bind the DnaA protein to initiate bacterial DNA replication
HU and IHF also bind
This causes the region to wrap around the DnaA proteins and separates the AT-rich region
HU and IHF
Composed of six subunits, first to come in into initiation of bacterial DNA replication
Helicase
Helicase travels along the DNA in the ___ direction, ___of polymerase
5’ to 3’, opposite
Helicase uses what to break the hydrogen bonds between the DNA strands
ATP
Separates the DNA in both directions, creating 2 replication forks
DNA helicase
The synthesis of leading and lagging strands from a…
single origin of replication
Polymerases connected in what strands
Leading and lagging
Topoisomerase is located where on the helicase
Head of helicase
Role of the single stranded binding proteins?
Keep the parental strands apart, slows down the re-annealing of DNA strands
Synthesizes an RNA primer
Primase
Synthesizes a daughter strand of DNA
DNA poly 3
Excises the RNA primers and fills in with DNA
DNA polymerase I
Covalently links the Okazaki fragments together
DNA ligase
The unwinding of DNA by ___ generates positive supercoiling ahead of each replication fork but ___ (a.k.a. topoisomerase II) travels ahead of the helicase and alleviates these supercoils
DNA helicase, DNA gyrase
bind to the separated DNA strands to keep them apart
Single-strand binding proteins
Short RNA primers are synthesized by…
DNA primase
Keeps genetic material from accumulating mutations at a high rate
RNA primers
The leading strand has how many primers, the lagging strand has how many primers?
Leading strand- single primer
Lagging strand- mutiple primers
Extends from 1 RNA primer and when it reaches the next primer—comes off and gets placed by DNA polymerase I
DNA poly 3
RNA primer on leading strand is replaced by ___
DNA poly 3 after 11 bases
attaches nucleotides in a 5’ to 3’ direction as it slides
toward the opening of the replication in ___
dna poly 3
in what strand: the direction is away from the replication fork
lagging
What polyermase uses the RNA primers to synthesize small
DNA fragments (1000 to 2000 nucleotides each)? These are termed?
3, okazaki fragments
DNA poly is in __ direction in lagging strand
opposite
Helicase is ___ of polyermase direction
opposite
If wrong nucleotide gets base paired and ligated into DNA, this will remove it
3’ exonuclease site
What DNA polys cannot initiate DNA synthesis? What does this instead?
DNA 1 AND 3, RNA PRIMASE
What DNA poly: Responsible for subsequent
DNA replication until it meets RNA
3
Takes over and removes the
RNA primers using its 5’ to 3’ exonuclease
activity and replaces them with DNA
DNA poly 1
Note that both Pol III and Pol I can only
attach nucleotides only in the ___
direction
5’ to 3’
links two adjacent single DNA
strands together
DNA ligase
The two copies of DNA Poly ___
are attached to each other
3
unlinks the circular daughter
chromosomes at the termination of Bacterial Replication
topoisomerase
Proofreading function
by DNA polymerases
involve
‘exonuclease
sites’
At what rate can bacterial cells divide into 2 daughter cells?
Very fast (faster than eukaryotes)
E.coli division rate
20-30 mins
Why is DNA replication in eukaryotes is more
complex?
large linear chromosomes, tight packaging with nucleosomes, more complicated regulation
How many origins of replication in eukaryotes
mutiple to ensure that DNA can be replicated in time
provided evidence for the multiple origins of
replication
Huberman and Riggs
How did Huberman and Riggs provide evidence for the multiple origins of
replication
They fed growing eukaryotic cells radioactive deoxythymidine that got
incorporated into DNA and then exposed DNA to film
Huberman and Arthur Riggs used a clever ____
experiment to show that mammalian cells are replicated by numerous
bidirectional origins
thymidine labeling
Huberman and riggs found
more than 2 replication forks (s phase)
what has a strong influence when origins get replicated?
chromatin
In contrast to bacteria, the leading and lagging strands (in regards to polymerases) are replicated by ___ in eukaryotes
different polymerases
In bacteria, the ___ is involved in both the leading and lagging strands and
___ helps out out on the lagging strands
dna poly 3, dna poly 1
In eukaryotes, pol ___ does the the leading strand while pol ___ and pol ___ do the lagging strand
poly epsilon, poly alpha, poly delta
Bacteria or eukaryotes: the polymerase taking over from primase (pol III) synthesizes most of the lagging strands before being replaced (by pol I)
Bacteria
In eukaryotes: the polymerase taking over from primase (pol ___) is in a complex with primase and synthesizes only a little bit before being replaced by pol ___.
alpha, delta (s shape)
How do the ends of linear eukaryotic
chromosomes get replicated?
have repepitive sequences at their ends (most telomeres have these) TTAGGC
In eukaryotes, DNA polymerase ___ link two nucleotides togethers ___ a primer
cannot, without
Ends of eukaryotic linear
chromosomes are
replicated by an enzyme
called
telomerase
Adds on DNA repeats using an RNA template to ends. RNA template has 1.5 copied of telomere
ribozyme
The telomeric repeat in eukaryotic chromosomes have how many bases
6
site for RNA polymerase
binding; signals the beginning of
transcription.
Promoter
site for the binding of
regulatory proteins; the role of regulatory
proteins is to influence the rate of transcription.
In eukaryotes, regulatory sequences can be
found in a variety of locations.
Regulatory sequences
Parts of DNA sequence for transcription
promotor, regulatory sequence, terminator
translation begins near this site in the mRNA. In
eukaryotes, a ribosome scans the mRNA for a
start codon
ribosomal binding site
specifies the first amino acid in a
protein sequence, usually a formylmethionine
(in bacteria) or a methionine (in eukaryotes)
start codon
Link AA together from start codon to stop codon
ribosomal binding site
a 3-nucleotide sequence within the mRNA
that specifies a particular amino acid.
codons
The sequence of ___ within mRNA determines the
sequence of amino acids within a polypeptide
codons
Bacterial mRNA may be ___, which
means it encodes two or more polypeptides.
polycistronic
What is a closed complex
when polymerase is bound to promotor and strands haven’t separated yet
The ___ functions as a recognition site for transcription factors
promoter
The transcription factor(s) enables ___ to
bind to the promoter. Following binding, the DNA is denatured into a
bubble known as the ___.
RNA polymerase, open complex
RNA polymerase slides along the DNA in an ___ to synthesize RNA
open
complex
causes RNA
polymerase and the RNA transcript
to dissociate from the DNA
termination signal
Prokaryotes have ____ RNA polymease, eukaryotes have ___
1, 3
A large proportion of bacterial promotors are organized like this, with….
-10 and -35 sequences conserved and 1st base pair with rna nucleotide layed down
In bacterial promoters, what is relationship with consensus sequences and promoters
the closer to the Consensus Sequence, the stronger the promoter
In bacterial promoters, when there is a weak conformity to consensus sequence what happens to transcription
less
In bacterial promoters, -10 sequence is known as a
pribnow box
In the initiation of bacterial transcription, Binding of RNA polymerase holoenzyme forms a…
closed complex
Describe bacterial transcription
- RNA poly core enzyme moves along DNA until they reach a promotor.
- Sigma factor then binds -10 and -35, stopping RNA polymerase from moving on DNA
- When is stops, RNA polymerase separates 2 DNA strands, initiating RNA synthesis and creating an open complex.
- As RNA initiates synthesis, a conformational change in RNA poly occurs and lets go of sigma factor
- When RNA poly releases sigma factor, theres a shape change in sigma factor and it lets go of -10 and -35 sequences
Many bacterial transcripts are terminated using the protein called…
rho
