human genome and mutation detection part1/2 Flashcards
how much nuclear DNA codes for functional gene products
2.5%
where do you find polymorphic DNA sequences
in extragenic sequences
what are polymorphic sequences closely linked to
linked to defined markers like gene, PCR primer. can also be linked to mutated genes.
what are minisatellite sequences used for
to determine paternity and to map mutant genes and detect individuals
what is another name for minsatellite sequences
VNTR variable number of tandem repeats
how can you detect VNTRs
by multi locus probes detect different but related sequences.
single locus probes identify one specific sequence
what does DNA profiling target
STR short tandem repeat
how do you amplify STR sequence
by PCR
how are PCR products analysed
by capillary electrophoresis
what is LCN low copy number of amplification
it amplifies DNA from a very small cell numbers
name a type of indirect DNA detection
by linkage analysis. You link polymorphic DNA markers with disease.
what can direct detection do
can detect deletions, insertions by PCR or southern blotting. can detect point mutations.
give examples of diseases caused by point mutations
sickle cell haemoglobin
alpha1-antitrypsin
give examples of diseases caused by deletions
cystic fibrosis
muscular dystrophy
thalassaemia
give examples of diseases caused by insertions
LDL receptor hypercholesterolaemia
how does allele specific oligonucleotides work
you need to define hybridisation and washing conditions precisely. need to label the oligonucleotide.
what is ARMS
amplification refractory mutation system
name some detection systems
1)specific detection of amplified products-Taqman, molecular beacons
2)non specific - SYBRG in real time PCR
3) fluoresence transfer and chemiluminescences system
4) scorpion probes
how can you detect known mutations
by allele-specific oligonucleotides hybridisation, PCR, gene chip
how do you detect unknown mutations
by DNA sequencing
what is first generation sequencing
shotgun sequencing- done by synthesis. High accuracy. Long read lengths. small data generated. example: ABI capillary sequencer
what is second generation sequencing
massively parallel sequencing- by synthesis, less starting material needed, high accuracy, short read lenghts. example; MiSeq (illumina) Ion Torrent
what is 3rd generation
single molecule sequencing- isngle molecule templates, low accuracy, long read lengths. example : single molecule real time sequencing. MinION
