Which alignment method and indexing options are suitable to use with Sierra? about sierra

[davhum]

Hi @hkarakurt8742,

We have only really tested Sierra on pipelines that utilise the STAR aligner (eg cellranger or STARsolo). In this case it is best to incorporate splice junctions via gtf when building the genome index.

I don't believe Hisat2 can be used for 10x single cell data - so you may want to double check this. Note Hisat2 can be used for full length single cell - but this isn't the kind of data Sierra was designed to work on.

[hkarakurt8742]

Hi @hkarakurt8742,

We have only really tested Sierra on pipelines that utilise the STAR aligner (eg cellranger or STARsolo). In this case it is best to incorporate splice junctions via gtf when building the genome index.

I don't believe Hisat2 can be used for 10x single cell data - so you may want to double check this. Note Hisat2 can be used for full length single cell - but this isn't the kind of data Sierra was designed to work on.

Thank you for your answer. I will keep working with STAR.
I just have a question about your answer. You mentioned that Sierra was not designed to work on full length transcript single cell RNA-Seq data so, is it not suitable to use a Smart-Seq2 based scRNA-Seq data with Sierra?

[davhum]

Correct: Sierra is not suitable for smart-seq2 data. Reason being is that smart-seq2 provides coverage across the whole transcript and if there is enough coverage you should be able to determine transcript isoforms either visually or perhaps using tools designed for bulk RNA-Seq (although not sure how well these will perform). Reason Sierra doesn't work on smart-seq2 data is because it will not reliably find piles of reads (i.e. peaks) at consistent places within transcripts.