High-throughput RNA isoform sequencing using programmed cDNA concatenation

Aziz M Al'Khafaji; Jonathan T Smith; Kiran V Garimella; Mehrtash Babadi; Victoria Popic; Moshe Sade-Feldman; Michael Gatzen; Siranush Sarkizova; Marc A Schwartz; Emily M Blaum; Allyson Day; Maura Costello; Tera Bowers; Stacey Gabriel; Eric Banks; Anthony A Philippakis; Genevieve M Boland; Paul C Blainey; Nir Hacohen

doi:10.1038/s41587-023-01815-7

High-throughput RNA isoform sequencing using programmed cDNA concatenation

Nat Biotechnol. 2024 Apr;42(4):582-586. doi: 10.1038/s41587-023-01815-7. Epub 2023 Jun 8.

Authors

Aziz M Al'Khafaji^#¹, Jonathan T Smith^#², Kiran V Garimella^#³, Mehrtash Babadi^#⁴, Victoria Popic^#⁵, Moshe Sade-Feldman^{2

6}, Michael Gatzen², Siranush Sarkizova², Marc A Schwartz^{2

7

8

9}, Emily M Blaum^{2

6}, Allyson Day², Maura Costello², Tera Bowers², Stacey Gabriel², Eric Banks², Anthony A Philippakis², Genevieve M Boland¹⁰, Paul C Blainey^{11

12

13}, Nir Hacohen^{14

15

16

17}

Affiliations

¹ Broad Institute of MIT and Harvard, Cambridge, MA, USA. aalkhafa@broadinstitute.org.
² Broad Institute of MIT and Harvard, Cambridge, MA, USA.
³ Broad Institute of MIT and Harvard, Cambridge, MA, USA. kiran@broadinstitute.org.
⁴ Broad Institute of MIT and Harvard, Cambridge, MA, USA. mehrtash@broadinstitute.org.
⁵ Broad Institute of MIT and Harvard, Cambridge, MA, USA. vpopic@broadinstitute.org.
⁶ Department of Medicine, Center for Cancer Research, Massachusetts General Hospital, Boston, MA, USA.
⁷ Department of Pediatrics, Harvard Medical School, Boston, MA, USA.
⁸ Division of Hematology/Oncology, Boston Children's Hospital, Boston, MA, USA.
⁹ Department of Pediatric Oncology, Dana Farber Cancer Institute, Boston, MA, USA.
¹⁰ Division of Surgical Oncology, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.
¹¹ Broad Institute of MIT and Harvard, Cambridge, MA, USA. pblainey@broadinstitute.org.
¹² Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. pblainey@broadinstitute.org.
¹³ Koch Institute for Integrative Cancer Research at the Massachusetts Institute of Technology, Cambridge, MA, USA. pblainey@broadinstitute.org.
¹⁴ Broad Institute of MIT and Harvard, Cambridge, MA, USA. nhacohen@broadinstitute.org.
¹⁵ Department of Medicine, Center for Cancer Research, Massachusetts General Hospital, Boston, MA, USA. nhacohen@broadinstitute.org.
¹⁶ Harvard Medical School, Boston, MA, USA. nhacohen@broadinstitute.org.
¹⁷ Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Charlestown, MA, USA. nhacohen@broadinstitute.org.

^# Contributed equally.

PMID: 37291427
DOI: 10.1038/s41587-023-01815-7

Abstract

Full-length RNA-sequencing methods using long-read technologies can capture complete transcript isoforms, but their throughput is limited. We introduce multiplexed arrays isoform sequencing (MAS-ISO-seq), a technique for programmably concatenating complementary DNAs (cDNAs) into molecules optimal for long-read sequencing, increasing the throughput >15-fold to nearly 40 million cDNA reads per run on the Sequel IIe sequencer. When applied to single-cell RNA sequencing of tumor-infiltrating T cells, MAS-ISO-seq demonstrated a 12- to 32-fold increase in the discovery of differentially spliced genes.

MeSH terms

DNA, Complementary / genetics
Gene Expression Profiling / methods
High-Throughput Nucleotide Sequencing* / methods
Protein Isoforms / genetics
RNA / genetics
RNA Isoforms* / genetics
Sequence Analysis, RNA / methods
Transcriptome

Substances

DNA, Complementary
RNA Isoforms
Protein Isoforms
RNA

Abstract

MeSH terms

Substances

Grants and funding