A universal and cost-efficient sample labeling approach for multiplexed single-cell RNA-seq based on recombinant...

Recent advances in single-cell transcriptomics have revolutionized our understanding of cellular diversity and tissue heterogeneity, providing unprecedented insights into biological and medical research. However, the high per-assay cost limits broader applications of this technology. Although sample labeling strategies enabling multiplexing have emerged, current methods suffer from either impractical complexity for barcoding or high cost for preparing the index labeling reagents. To address these challenges, here we present HUH-endonuclease-agglutinin tagging (HEATag), a universal cell membrane labeling approach that combines Duck circovirus HUH endonuclease (DCV) with wheat germ agglutinin (WGA) to efficiently tag cell membranes with indexed single-stranded DNA (ssDNA). The DCV domain enables rapid, sequence-specific conjugation of indexed ssDNA, while the WGA domain ensures robust labeling of fresh or fixed cells across diverse species. This method is compatible with both commercial platforms and custom systems, readily adaptable to various single-cell omics workflows. Therefore, HEATag provides a universal, cost-effective, and scalable solution for high-throughput single-cell studies, enhancing library preparation efficiency and minimizing batch effects for single-cell researchers.