SNP genotyping, copy number variation, gene expression levels etc. In each case you know what you are looking for and in fact could do it with PCR but if you have lots of samples, you may choose to do NGS for cost and turnaround time.
Ever heard of 16S sequencing? There are parts of the 16S gene that are conserved, and parts that are hypervariable. The hypervariable region (in a perfect world) is the same for all members of a single species but are different between species (i.e. all members of bacteria species 1 will have the same hypervariable region sequence, and all members of bacteria species 2 will all have the same hypervariable region sequence, but the sequence in bacteria species 1 will be different from bacteria species 2). The conserved region (again, in a perfect world), will be the same across ALL species. Because the sequences between species are unique, when we sequence this region, we can identify species based off the hypervariable sequence. Because we know the sequence of the conserved region, and that the conserved regions from bacteria species 1 and 2 will be the same, we can use the same primers to amplify the target gene from either species, creating amplicons. One can then sequence these amplicons to figure out whether bacteria species 1 and/or 2 is in a sample.
My grandpa was a gravity module maintenance guy all his life. He used to say to me all the time "You gotta pay the bill if you wanna stay still". Not a great slogan, but that's why Pop Pop was a maintenance guy and not an exec.
Yes, the adapters used for Ion library preparation are specific to Ion platforms. This is because we use specific primer sequences which are complementary to the adapters, during template preparation and sequencing.
Natalie explains it so well! Great posts so far.
1:27 why would someone want to sequence DNA with an allready known sequence?
Exactly my question...
SNP genotyping, copy number variation, gene expression levels etc. In each case you know what you are looking for and in fact could do it with PCR but if you have lots of samples, you may choose to do NGS for cost and turnaround time.
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to look for variations between samples of a known sequence for instance
Ever heard of 16S sequencing? There are parts of the 16S gene that are conserved, and parts that are hypervariable. The hypervariable region (in a perfect world) is the same for all members of a single species but are different between species (i.e. all members of bacteria species 1 will have the same hypervariable region sequence, and all members of bacteria species 2 will all have the same hypervariable region sequence, but the sequence in bacteria species 1 will be different from bacteria species 2). The conserved region (again, in a perfect world), will be the same across ALL species. Because the sequences between species are unique, when we sequence this region, we can identify species based off the hypervariable sequence. Because we know the sequence of the conserved region, and that the conserved regions from bacteria species 1 and 2 will be the same, we can use the same primers to amplify the target gene from either species, creating amplicons. One can then sequence these amplicons to figure out whether bacteria species 1 and/or 2 is in a sample.
really easy to understand and interesting, thanks!
Great overview! Would you mind explaining how size selection impacts efficiency and quality of downstream sequencing?
Great video!
Can you explain how to read NGS data
Thank you so much maam very helpful information for bigginers
Natalie you are awsome
gold pipette
Was it a reference to Austin Powers? haha
So helpful!
which adapter attaches to the bead?
The barcode adapter, or the P1-Adapter?
Hi Barbara. Yes, the P1 adapter on the library fragments anneal to the ISPs, and the complementary strand extends from the ISP during amplification.
I forgot to go pay our gravity bill. Gotta run turtles
My grandpa was a gravity module maintenance guy all his life. He used to say to me all the time "You gotta pay the bill if you wanna stay still". Not a great slogan, but that's why Pop Pop was a maintenance guy and not an exec.
Are the adapters sequencing-platform dependent?
Yes, the adapters used for Ion library preparation are specific to Ion platforms. This is because we use specific primer sequences which are complementary to the adapters, during template preparation and sequencing.
Thermo Fisher Scientific Thanks for the reply.
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Great video!!!