Thread by @MackayIM

@satchitanand21 @ColleenHuberNMD So yeah, the thing is the human genome is big & has bits & pieces that sometimes match up with bits & pieces in other [parts of the tree of life. But that doesn't really mean anything by itself. Unless you're doing a PhD on the topic I guess.
So I took that primer sequence...

@satchitanand21 @ColleenHuberNMD ..it's a reverse primer (so actually hybridises to the reverse complement of that sequence; long story I won't cover here if you didn't get what I just said).
If we take that sequence and compare it against a tree-of-life sequence database (called BLAST-blast.ncbi.nlm.nih.gov/Blast.cgi)..

@satchitanand21 @ColleenHuberNMD ...as Capt Conspiracy did...we get some results.
First up I've excluded any matches to SARS-CoV-2, there is indeed 100% match to human DNA but also dog, chimp, malaria parasites, some bacteria, fish & bird. Guess what?

@satchitanand21 @ColleenHuberNMD This is actually pretty much normal ho-hum level stuff when you look at one short (oligo) sequence in isolation. Any PCR designer knows this like they know which shoe to put on which foot.
Capt Conspiracy may not be much of an expert.
Sorry.
But I think you need to know this.

@satchitanand21 @ColleenHuberNMD But what about if I allow the inclusion of matches to SARS-CoV-2 - the intended target of this French real-time RT-PCR test - I hear you ask?
Thank you.
That's a sensible question.
Much moreso than your first one.

@satchitanand21 @ColleenHuberNMD Lo and behold, matches to SARS-CoV-2 fill up the screen. Hooraaaaaaaaayy!

@satchitanand21 @ColleenHuberNMD But what about the human DNA match, you ask next??
Well, it turns out that for PCR to work, it needs *two* primers to work.
I've written a little bit on this (needs improving but might give you the gist)'
virologydownunder.com/pcr-primers-a-…

@satchitanand21 @ColleenHuberNMD And for real-time RT-PCR - which this is one of - there is also a probe - so 3 distinct short sequences all have to bind for the thing to work at all.
Otherwise - it doesn't work.
At all.
Did Capt. Conspiracy talk about that? Hmm? Nope.
#failCapt

@satchitanand21 @ColleenHuberNMD Yeah, I know what you're thinking. What happens if you BLAST the other (forward) primer & the probe for this French SARS-CoV-2 real-time-RT-PCR?
Another great question buddy.

@satchitanand21 @ColleenHuberNMD Turns out, they don't match that human DNA so much.
But they also match really well to SARS-CoV-2 RNA.
So think of this as a torch-without a battery, bulb & circuit it won't work. Put them together, it's a torch.
Yes, the expert knew WTF they were doing. Phew. Just lucky I guess/