Best Pipeline for De novo transcriptome SC-RNAseq

Question

[Chris]

Hi everyone,

I'm curious what the best pipeline for single cell RNA seq is if your organism of interest doesn't have a genome. Tools such as CellRanger need a reference genome, and bustools seems to need a reference for indexing.

Is the best approach to trim the UMIs from the files and perform a de novo assembly using a tool like trinity, then use this as the reference genome for CellRanger / Bustools?

Many thanks,

Chris

[ATpoint]

I doubt that the sparseness of single-cell data allow any quality assembly. The least you need is a reference transcriptome, not necessarily a genome. Tools like salmon-alevin or mentioned kallisto-bustools can quantify directly against a transcriptome.

[dsull]

You can try bulking the single-cell data together and do a transcriptome assembly from the RNA-seq reads.

Better yet, do bulk RNA-seq on your organism and do a Trinity assembly that way.