Moelcular Medicine I Flashcards
what the 2 phases of genetic analysis of a disease
find a correlation between amuattion an da diseae
detect or predicting a disease
when is DNA sequencing most useful
where the exact nature of the disase causing muation is known
limits to DNA sequencing
genes are large and complex, some of the mautions may not be connected to the disease at all
indirect methods advantage
require no knowledge about the exact nature of the muation as long as the genomic locus of the mutant allele is known
limits to indirect methods
good polymorphic markers are scarce. and you could not be looking in the right place
the complete human genome consists of how many basepairs
3 billion
WGS
looking at the whole genome
WES
only looks at the exons
SNP typing
looking just at SNPs which are the areas that are actually different between people
wha tis the narrowest approach to sequcning
just look at the SNPs that have been assocaited with a health risk
limiations to seuqncing
- doesn’t show on which chromsome the muation is
- does not show if they have a deletion of a duplicatio nof a chromsomal region
copy number variation (CNV)
observation that chromsomal regions can be present in non-diploid states
most CNVs are what
benign polymorphisms
how do you do comparative genome hybridization
take sample of single stranded genomic DNA from pt and control. label with die, mybrize to metaphase chromosomes. CNV becomes evident by uneven labeling of template chromsome
FISH
allows ID of a chromosomal lobus on a metaphase chromosome by hybridation with a specific flouorescent probe
how do you do PCR
- denature template DNA
- annel primers
- polymerase synthesizes complmentary strand
how does PCR detect genetic polymorphisms
when using ASOs, only mutant alleles will be amplied
insertions or deletions become immediately obvious
how do you do real time PCR
- the amoun tof amplified DNA is measured after each cycle
- determine how many cycles of amplification are required
- high number of templant molecules, fewer cycles will be required to reach the threshold
- templant amount is quantified by the number of cycles required to reach threshold
what is reverse transcriptase PCR used for
retroviral load in blood samples (HIV)
expression of oncogenes, and tumor suppressor genes in cancer cells
ASOs
tool to detect single nucletotide polymorphisms and mutations
what does ASP require
detailed knowledge about the nucletodie sequence of the gene under studyy
because oligonucleotide primers are short they anneal only to gerions of ______
high sequence complmentarity
mutant allele in aickle cell
AT exchange in codon 6 of beta globin gene
DNA microarrays
thousands of oligonucletodie spots printed on an inch paper. use for genetic screening
the differences in individual drug responses result from what
genetic polymorphisms
goal of pharmacogeomics
find the polymorphisms taht affect drug responses in order to work out an indvidual drug treatment plan
what is malignant hyperthermia
rare, authosomal disoder that is the leading cause of deal during anesthesia
what happens when someone with malignant hyperthermia goes under anthestia
calcium gets released from muscle cells leading to life threatening episodes of uncontrolled muscel contraction
what causes malignant hyperthermia
mutation in ryanodine-receptor (RYR1) gene coding fo rmuscel cell membrane ion channel
individual response to warfarin is determined by what 2 enzymes
- gene for detoxifying P450 protein causing slow metabolism of drug
- mutation in vitamin K epoxide reductase
polymorphism in gene for detoxifying P450 protein causes what
higher senstivity to warfarin
mutation in vitamin K epoxide reductase complex (VKORC1) leads to waht
higher warfarin toelrance
how are fetal cells sampled
amniocentesis
chorionic villus sampling
amniocentesis
amniotic fluid is withdrawn at 15/16 weeks gestation
chorionic villus sampling
cells taken from chorion at 10-12 weeks gestation
what condition might chonrionic villus sampling miss
mosaicism
the specific bindign of antibodies to antigens is used to what
- detect presence of antigen in sample
- quantify amount of antigen in sample
- localize antigens in vivo
what produces antiboties
b lymphocytes
how are polyclonal antibodies obtained
challenged an animal with intravenously injected antigen. , then after several days take blood
how do you get monoclonal antiodies
- inject mouse with protein
- get spleen cells
- fuse spleen celsl with myeloma cells grown in culture
- select and grow hybrid cells
- select and grow cells making desired antibody
hybridoma cells
immortal and produce antibodies
what is ELISA commonly used for
detecting PTH
HbA1c
c-reactive protein
what do you use sandwich ELISA for
quantify an antigen
What do you use indirect ELISA for
presence of antibodies
What specific diseases is indirect ELISA used for
HIV
rheumatoid arthritis
western blotting
technique that uses specific antibodies to indicate the molecular weight of a protein
what is the advantage of western blotting over ELISA
you get better infromation about the antigen
what do you use western blotting for
confirm diagnosis of HIV
diagnose parasite infections
