Using CB_UMI_Complex for paired-end reads

[Josephine Bagnall]

Hi,

I'm trying to use STAR solo where there is UMI + barcode1 + barcode2  + cDNA in Read2. I could specify these barcode locations using CB_UMI_complex, but then I received an error when trying to specify that soloBarcodeMate was 2 (the barcodes are on read2). It said that I would need to have the barcodes on a separate file (soloBarcodeMate 0). However, the documentation for CB_UMI_complex states that it expects barcodes on Read2.

If I separate out the barcodes to it's own file, I could specify three input files (readFilesIn) to be Read1(cDNA only)  Read2(cDNA only) Barcodes(UMI+barcode1+barcode2). If I do this, will STAR solo use Read2 at all to do the paired end alignment? or will it ignore Read2 and use Read1 only for single end alignment?

Could you please advise the best way to do paired-end alignment with UMI & 2 barcodes?

Thanks for the help,

Josephine

[Alexander Dobin]

Hi Josephine,

Presently, --soloBarcodeMate only works with CB_UMI_simple.

If you use CB_UMI_complex, it will need to discard the cDNA on Read2.

The workaround is to concatenate barcode1 and 2 into a single sequence and create a combined passlist.

[Josephine Bagnall]

Hi Alex,

Thank you for the clarification!

Best,

Josephine