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bioRxiv · 10.1101/2025.02.19.638981

Spatial Total RNA Sequencing of Formalin-Fixed Paraffin Embedded Tissues by spRandom-seq

Liao, Y.·Chen, J.·Zhang, S.·Chen, Y.-s.·Zhu, B.·Chen, H.·Zhu, Y.·Xu, Z.·Yin, Y.·Xiong, X.·Liu, N.·Guo, H.·Wang, Y.
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Abstract

Current oligo(dT) primers-based spatial transcriptomic methods are limited to fresh or fresh-frozen samples due to the low efficiency of oligo(dT) primers in capturing RNAs in degraded or microbial samples. Here, we have developed a random primer-based spatial total RNA sequencing (spRandom-seq) technology for simultaneously capturing whole host and microbial RNAs in formalin-fixed paraffin-embedded (FFPE) tissues. spRandom-seq eliminated 3 or 5 gene-body biases and outperformed oligo(dT)-based 10X Visium with an 8-fold higher capturing rate for lncRNA and other non-polyadenylated RNA biotypes, including miRNA, snRNA and miscRNA. In the clinical FFPE sections of breast cancer, we revealed the inherent heterogeneity within the tumor region. We also simultaneously captured host and microbial RNAs in Klebsiella pneumoniae-infected sections. Totally, spRandom-seq provided a versatile tool for both clinical pathology and infection biology with established spatial platforms, ensuring ease of operation and large-scale applications.

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Liao, Y., Chen, J., Zhang, S., Chen, Y.-s., Zhu, B., Chen, H., Zhu, Y., Xu, Z., Yin, Y., Xiong, X., Liu, N., Guo, H., Wang, Y.. 2025-02-23. Spatial Total RNA Sequencing of Formalin-Fixed Paraffin Embedded Tissues by spRandom-seq. https://doi.org/10.1101/2025.02.19.638981

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