Genetic Testing Flashcards
(31 cards)
how does sanger sequencing fragment DNA
flourescent terminating nucleotides
what makes sanger harder to interpret
shift mutations- insertions/ deletions
what can NGS do
sequence many regions of DNA at the same time as samples spatially separated
millions of overlapping DNA fragments generated for each sample (lots of data)
barcode identifies specific samples (demultiplexing)
in NGS what is a paired read
when fragment is sequenced from both ends
how long is a short read in NGS
300bp
what does coverage mean
proportion of genes covered by testing
depth of coverage= how many times each nucleotide is sequenced
(for germline want coverage of .95% at x30 depth)
(for somatic x200 depth)
uniform coverage of a sequence means you have to do it less as more areas covered well
what is NGS most commonly used for
gene panels
what does the overlap of fragments allow
re-ordering
what is a duplicate read
when the same region sequenced twice- waste of time
what are the disadvantages of short read NGS
GC bias
15 hours +
not good at detecting transcript splicing variants or structural DNA variants (translocations/ fusions) as hard to get sequence that covers the break/ splice site
what does long read NGS allow
real time sequencing
no GC bias
dont need to amplify DNA
direct detection of modifications such as methylation
is short or long read sequencing used for wgs
short
why does wgs variant interpretation take a long time
as variants of unknown significance identified
what does targeted sequencing via hybridisation allow
sequencing clinical exome, whole exome or specific genes
cheaper, less unknown variants
what can targeted sequencing via PCR do
amplify regions of interest
useful where limited sample available e.g. cancer biopsy
what can targeted long read sequencing do
chromosomal rearrangements
what is consituitive genetics
gene panel
where small number of genes associated with phenotye
what is chip-seq
assesses DNA binding proteins
what is atac-seq
unwound sequences- transcriptionally active
what is RNA sequencing good for
fusion transcripts (formed from chromo rearrangements) used in diagnostic cancer testing
what is cDNA
DNA formed from RNA by reverse transcriptase
what do microarrays do
by comparison to reference show variants present
methylation assays
SNP assays - only detect known possible variants
can only detect unbalances abnormalities (microdeletions/ microduplications, chromo abnormalities, excessive homozygosity)
what is the difference between genetic and genomic testing
genetic= changes in individual genes/ proteins genomic= changes at multi gene or structural level
what is cancer germline testing
inherited cancer syndromes
