Mapping file

[Lekha Menon]

Hi,

I am creating my mapping file and I have a doubt. I have the Fusion (Linker) +Key+Barcode+Primer and then in the reverse primer I have Fusion (Linker)+Key+Primer. So does the linker primer sequence mean the entire one without the barcode or just the Linker alone? Please help. 

Thanks,

Lekha

[Jai Ram Rideout]

Hi Lekha,

I've contacted a developer to help answer your question.

Best,

Jai

[TonyWalters]

Hello,

The linker you are describing is different than the linker that is part of the LinkerPrimerSequence value. It usually is just 2 nucleotides, and sits between the barcode and the primer. I would just put everything after (3' direction) the barcode portion of your construct under the LinkerPrimerSequence field. For the ReversePrimer, I would just do the primer component.

-Tony

[TonyWalters]

Hello Lekha,

That should be fine, since this line is zero:
Percent of sequences with barcodes detected at the beginning of the sequence: 0.000
It looks like there are some small fraction of the reads that happen to have one of the sequences matching your barcode *somewhere* within the read itself, which can happen with short sequences like the barcode.