Can RNA data generated from the multiome kit be analyzed using the scFAST-seq pipeline (SeekSoul Tools fast module)?

Answer:
Yes, but this is generally not recommended as the primary analysis strategy due to differences in sequencing quality characteristics.

The RNA library structure generated by the multiome workflow is fundamentally identical to the scFAST-seq workflow, because both use the same gel bead semi-random priming design. Therefore, in principle, multiome RNA libraries can be analyzed using the SeekSoul Tools fast module.

However, an important difference is sequencing context. In multiome experiments, RNA libraries are sequenced together with ATAC libraries. Because ATAC libraries contain relatively low-diversity R1 sequences, the R1 quality of RNA libraries may be somewhat lower than in standalone full-length transcriptome sequencing runs.

The SeekSoul Tools fast module performs alignment using both R1 and R2 reads. Consequently:

• Lower-quality R1 bases may reduce overall alignment rates 
• Additional mismatches or alignment artifacts may be introduced in some datasets 

For this reason, the SeekArc Tools RNA module is generally recommended as the default analysis workflow for multiome-generated RNA data.