Mod 3 Genomes Flashcards
what is a genome
the complete set of DNA molecules that an organism posses
how many bP of DNA in human genome
+ detail
3200 Mb of DNA
split into 24 linear DNA molecules
shortest is 48 Mb
longest is 250 Mb
how many chromosomes in a normal diplod cell?
6400 Mb
22 sets of chromosomes + either XX or XY
how many bP in e.coli genome
4.64Mb
in a single DNA (circular double stranded DNA)
describe the replication of the E coli genome
begins at origin of replication
always the same position on the genome and only one origin
two replication forks (bidirectional - cuz circle so splits into two directions)
describe the replication of human DNA
origins of replication
many orgins on each chromosomal DNA molecules
each fork copies about 150kbof DNA (so has to be loads of origins)
how does the origin of replication form
barrel of DnaA proteins
the DNA strand wraps around this barrel of proteins
this forces the base pairs at the origin of rep to break
cuz they’re mainly A-T pairs (only 2 H bonds so are easier to break)
how does the pre priming complex form
DnaB proteins attach to origin of rep site
DnaB is a helicase
so this breaks more base pairs so the replication fork moves away from the origin (bidirectionally)
how is the primosome formed
two primase enzymes attach
these make RNA primers that initiate replication of the two leading strands
what order does the preparation for replication go in
origin of rep forms
pre priming complex forms
primosome forms
then events at replication fork begin as per usual
summary of replication process at the rep fork (prokaryotes)
(including gamma and beta complexes)
- helicase (DnaB) breaks base pairs
- Single Strand Binding proteins (SSBs) protect the bare single strands
- DNA topoisomerase unwinds strands and sorts out supercoiles
- Primase makes primers on leading and lagging strands
- DNA pol III synthesises DNA
- DNA pol I and DNA ligase removes primers and joins Okazaki fragments
- gamma complex (aka clamp loader) - attaches and detaches pol III from the lagging strand
- beta complex (sliding clamp) - holds pol III onto template, alowing it to slide
summary of replication process at the rep fork (eukaryotes)
- helicase (DnaB) breaks base pairs
- Single Strand Binding proteins (SSBs) protect the bare single strands
- DNA topoisomerase unwinds strands and sorts out supercoiles
- Primase and DNA pol a makes primers on leading and lagging strands
5.DNA pol a makes first 20 bp - DNA d takes over (two copies of DNA d) synthesises DNA
- FEN1 and DNA ligase removes primers and joins Okazaki fragments
- proliferating cell nuclear antigen (PCNA) (sliding clamp)
holds DNA pol delta onto DNA
what are the names of the sliding clamps in prokaryotes and eukaryotes
p= beta complex
e= proliferating cell nuclear antigen PCNA
holds the DNA pol III or DNA pol delta to the DNA
how is the replication preocess terminated? (ecoli)
terminator sequences - eah of which are binding sites for a Tus protein
they have a specific orientation which decides how the Tus proteins bind
Tus protein
has round side and flat side
round = permissive
flat = non-permissive
as in the rep fork can or can’t pass through
the orientation of these relies on the terminator sequences and THEIR orientation
see diagram one note, kinda looks like raajma
Tus lock domain
when a rep fork tries to pass through the non-permissive
theres a ‘reserved’ C nucleotide in the terminator sequence
so when it tries to pass through, its sequence opens up and the C gets trapped
causes rep fork arrest
basically it cant pass trhough anymore and that’s how the non perm side works
how is replication terminated in human DNA
it just kinda merges into each other
doesn’t need the exact control like in circular DNA (terminator sequences and tus proteins blah blah)
what is chromatin
DNA extracted from the nucleus
a DNA-protein complex (can be shown with digestion with an endonulease)
proteins are spaced at regular intervals along the DNA (around 200bp)
looks like ‘beads on string’ structure
describe the digestion of chromatin with an endonuclease
DNA wrapped around multiple protein complexes (histones)
(=chromatin)
add in endonuclease treatment
it will ‘cut’ the DNA between 1 or 2 of the proteins (so its still wrapped)
if you degrade the histones, the DNA fragments that are produced are all 200bp or multiples of 200
what are the proteins called in chromatin
histones, 5 diff types
what are the beads in a chromatin called
nucelosomes
how many proteins does one nucleosome have
8 proteins (histones)
2 each of H2a, H2b, H3 and H4
what is linker DNA and how much
link nucelosomes together
50-70bp
what is the linker histone
Histone H1
attaches outside the nucleosome
brings the linker DNA together
kinda like mini clasps in a bracelet between the beads
what is a chromatosome
nucelosome + DNA + linker histone
how much does the beads on a string structure reduce the length of DNA
1/6th
how much does the 30nm chromatin fibre reduce the length of DNA
1/7th
so 4.9cm molecule in a chromosome would be about 1.2mm in length after this
describe 30nm chromatin fibre structure
pulls together the beads on a string to make it more compact
nucleosomes stack with ‘non-neighbours’ (so N1 is on top of N3 not N2)
it zig zags and connected by straight DNA linker
formed into tetranucelosomal units
see diagram onenote
what is the imagin technique to see these structures
cryo electron microscopy
how is the 30nm nucleosome fibre stabilised
using H1 which is off axis and asymmetrically bound
interacts with linker DNA again
how is the 30nm nucleome structure oriented correctly
histone H4 acidic N-terminal tail
it interacts with the H2a/H2b acidic patch in the nucleosome
how is it possile to change the chromatin structure
via modification of histone proteins (dont need to know detail, just know that it’s possible)
what is a possible effect of nucleosome presence or modification
controls gene expression
different types of chromatin and their functions
euchromatin (light areas on a picture)
=contains active genes (mostly as 30nm fibre forms)
heterochromatin (dark areas)
more densly packed
contains inactive genes
- constitutive heterchromatic = always tightly packed in all cells (e.g. one of the X chromosomes in females in always inactive)
- facultative heterochromatin = sometimes packed away, other times in use - depends on the cell type
what is in nucelolus
contains ribsomal RNA
site of ribosomal biogenesis (where its made)
euchromatin and how is the shape of the nucleus maintained
DNA is in form of 30nm fibers or less compact
attached to the nucelar matrix and nuclear lamina (both formed from proteins),not just floating baout
= this maintains the shape of nucleus
what are diseases associated with nuclear matrix and nuclear lamina
mutations that affect the formation of nuc matrix or lamina:
- progeria (premature ageing) - bad nuclear lamina - ruins struc of nucleus
- downs sydrome - nuclear matrix
- huntingtons disease - nuc matrix
what is the highest level of compaction in the DNA
metaphase chromosome - the 30nm fibres is folded into loops
only found in dividing cells
describe structure of metaphase chromosome: centromere
has centromere in middle:
- holds the daughter chromosomes together
- contains special histones CENP-A insterad of H3
- attachment point for microtubules that pull the chromosome apart
describe structure of metaphase chromosome: telomere
protects the ends:
from exonuclease attack
and from being mistake for chromosome breaks and joined together by DNA repair mechanisms
what is a karyogram
metaphase chromosomes are stained
give a banding pattern - uxed to map position of genes
so all 22 chromoses plus X and Y
