Prenatal Genetics Flashcards
chromatin composed of …
DNA, histones, and other proteins
chromosomes are made of …
chromatin
chromatin plays an important role in…
DNA compaction as well as regulating replication, transcription, recombination, and chromosome segregation
nucleosome cores are joined by …
linker DNA
- 34 bp
- nucleosome consists of a histone octamer, 147 bp of DNA wrapped around the octamer and linker DNA
a nucleosome histone core is composed of ___ histones
8
each nucleosome histone cor contains two of the following histones:
H2A, H2B, H3, H4
nucleosome core DNA is comprised of ____ bpm which wrapped around the histone octamer
147
nucleosome functions
- facilitates compaction of ~200 bp of DNA
- facilitates compaction of 30 nm chromatin fibre
- template for chromatin enzymes which facilitate post-translational modifications
- PTM facilitates higher levels of compaction
what is a chromatosome?
consists of a nucleosome and a linker histone
histone protein H1 may bind to linker DNA and protect an additional 15 to 20 bp of DNA
H1 bind at the DNA entering and exiting the nucleosome
most common linker histone
H1
heterochromatin
eukaryotic chromatin that remains tightly compacted during interphase and is not transcribed
telomere
end of a chromosome composed of repeated DNA sequences and associated proteins
p arm =
petite; short arm
placed at top
q arm =
long arm
bottom
centromere
appears as a constriction in metaphase chromosomes
- composed of repeated sequences in the ara of the chromosome that attaches to the mitotic spindle
satellite
part of the end of a chromosome that is separated from the rest of the chromosome by a secondary constriction
metacentric
centromere located centrally, p and q arms are approx. same length
sub metacentric
centromere located off center
q arm slightly longer than p arm
acrocentric
cetromere located nearer one end of the chromosome; q arm significantly larger than p arm
telocentric
centromere is the distal end of the chromosome
no p arm
not normally found in humans
MLPA
multiplx ligation-dependent probe amplification
- propietary technique developed by MRC Holland
applications of MLPA
- includes detection of deletions, duplications, and aneuploidies
- more sensitive for detecting insertions(?) and deletions
MLPA Steps
denaturation
Probe hybridization
Ligation
PCR
Capillary electrophoresis
analysis
when does chromatin modification happen?
before transcription
which part of histones are available for chemical modification
histone tails (protrude outwards)
______________ of histone tails promotes loose chromatin structure that permits transcription
acetylation
- when nucleosomes highly acetylated = chromatin less compact = DNA accessible for transcription
Euchromatin
less condensed form of eukaryotic chromatin = available for transcription
this agent arrests cells at metaphase
colcemid
describe steps in Giemsa staining
- dip slides in trypsin (frees histones and other proteins associated with DNA so Giemsa can stain DNA)
- rinse in saline
- incubate in Giemsa for ~1min
- rinse with tap water
- dry using fan
- view with light microscope
longer dry = sharper bands
the most common method for staining chromosomes
Geimsa staining
G-banding
- Giemsa for chromosomes
- lighter areas = euchromatin
- dark regions = heterochromatin
- centromeres do NOT stain
What is G-banding used for
to identify chromosome abnormality and rearrangements in genetic diseases and cancers
T or F. in G-banding, resolution is related to sensitivity
T
degree of resolution also varies depending on mitotic stage
karyotype
an individual’s complete set of chromosomes
ex: 46,XX
aneuploidies
abnormal # of chromosmes
most not ocmpatible w life
balanced translocation
usually compatible w life; ppl dont even know they have it
chromosomes sometimes don’t line up easily; can be reciprocal or one piece attached to the other
at least one good copy of each gene so not detrimental
ex: chr 13 broke off and attached to chr 5
unbalanced translocation
translocation between chromosomes resulting in gain or loss of genetic material, likely resulting in a chromosomal condition
- usually occurs when balanced traslocation person has children
isochrome
unbalanced structural abnormality
- two p arms or two q arms separate together
- there’s both a loss and a gain of genetic material
what is FISH
deploys DNA probes specific for each chromosome (or subchromosomal region or single locus)
- probes fragments of DNA or RN = usually 100-1000bp
- used to detect presence of nucleotide sequences that are complementary to sequence in the probe
- probes are labeled with modified nucleotides that fluoresce under particular conditions
- by using different fluorochromes, a karyotype can be “painted” as desired
FISH whole chromosome probes
collections of smaller probes, each of which binds to a different sequence along length of a given chromosome
spectral karyotyping
FISH
- can label each chromosome a different colour
- VERY expensive so not used routinely
= 24 chromosme-specific painting probes are used i just one FISH experiment
FISH centromeric probes
- can be used on interphase and metaphase chromosomes
- used to enumerate chromosomes
- typically used as controls
locus specific FISH probes
bind to a particular region of a chromosome
- useful when scientists have isolaed a small portion of a gene and want to determine on which chromosome the gene is located or how many copies of a gene exists within a particular genome
> CNV probe, fusion probe, break-apart probe
G BANDING VS FISH
G-banding = large changes in chromosome
FISH = small changes not visible by G-banding or if we want fast results
CNV probes
designed to hybridize to a precise gene location
identify gene deletions and amplifications
gene fusion probes
occur when two normally separated genes are joined
while normal cell will display the colours as separate signals, a cell that has fusion will show the signals less than one signal width apart
break apart probes
a single probe w 2 fluorophores
probe target twoareas of a specific gene sequence
when intact= yellow
when apart = red and green seen
