Description

Spatial Isoform Profiling within Individual Tissue Sections

Michaela Asp¹, Alex Stuckey¹, Konstantin Carlberg¹, Joel Gruselius², Sebastian Johansson³, Erik Borgström², Fredrik Salmén¹, Sanja Vickovic¹, Max Käller¹, Patrik L. Ståhl¹, Joakim Lundeberg¹

¹Division of Gene Technology, KTH Royal Institute of Technology, Science for Life Laboratory, Stockholm, Sweden; ²Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden; ³Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden

Massively parallel cDNA sequencing has rapidly evolved from the traditional bulk analysis towards both single cell and spatially resolved gene expression analyses within tissue sections. This has broadened our understanding of biological systems, as bulk studies can only provide average measurements of gene activity. Today, individual single cells can be characterized in a high throughput manner and complete transcriptomes can be reconstructed. Spatial transcriptomics captures transcript tags and offers an efficient approach for quantitative gene expression studies, due to retaining spatial information about the tissue context. The tagged approach however restricts deeper investigations concerning alternative splicing, fusion transcripts and possible discoveries of new genes, which typically requires information from the full-length transcript to be present. Here we present a new strategy that allows a global view of spatially resolved full-length transcriptomes by using spatial transcriptomics positionally barcoded microarrays, in combination with full-length transcript information and cDNA sequencing.

Full-length cDNA from individual tissue sections are synthesized directly on the microarray surface based on the Smart-seq2 technology¹. Next, full-length cDNA with positional barcodes are released from the surface and amplified in a single pool. The cDNA library is then prepared so that the full-length information is retained when sequencing. By combing the positional barcodes from the microarray with the full-length constructs, we are then able to examine spatially resolved full-length transcripts for the determination of complete transcript structures.

1.Picelli, S. et al. Full-length RNA-seq from single cells using Smart-seq2. Nat. Protoc. 9, 171–81 (2014).