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Recombinase polymerase amplification to enable same-cell, DNA/RNA multi-omic single cell sequencing

Award Number: R21GM152950
ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 01/01/2024
PERIOD OF PERFORMANCE END DATE: 06/30/2026

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Recombinase polymerase amplification to enable same-cell, DNA/RNA multi-omic single cell sequencing - PROJECT SUMMARY/ABSTRACT: Single cell sequencing technologies have rapidly advanced over the past five years and have become essential tools in a broad array of fields, ranging from developmental biology to human genetics. One major advance was the development of high-throughput barcoding of RNA or DNA within droplets, which allowed for preparation of sequencing libraries as a pool rather than in a laborious 96 well plate format. This advance enabled sequencing analysis of thousands of cells in one batch. However, there has not been the same breakthrough in scale when sequencing both DNA and RNA from the same cell as established techniques involve sorting cells into plates or wells and ultimately separating DNA and RNA before library preparation. The separation of RNA from DNA is necessary because conventional PCR amplification requires a 95oC melting step, which degrades RNA. In this proposal, we will seek to optimize DNA amplification through recombination polymerase amplification (RPA), an isothermal form of PCR which can be performed simultaneously with reverse transcription of mRNA into cDNA. In this way, both RNA and DNA can be captured into barcoded, sequencing libraries within the same droplet. The technology could have impact in many different fields in which genotype-phenotype correlation is needed, such as characterizing genetic polymorphisms or genetic variants in cancer. In SA1, we will demonstrate proof-of-principle of same cell DNA amplicon and RNA transcriptome technology via isothermal RPA and reverse transcription. In SA2, we will determine the quantitative performance of the methodology in two use applications and test capability to genotype heterozygous alleles. If successful, this methodology could be widely adopted by laboratories in diverse fields to answer questions related to genotype-phenotype correlations.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2026 ( Subtotal = $0 )
2026	2025	SLOAN-KETTERING INSTITUTE FOR CANCER RESEARCH	1275 YORK AVE	NEW YORK	NY	10065	NEW YORK	USA	Biomedical Research and Research Training	000	2	3/13/2026	NON-COMPETING CONTINUATION	$0
														Subtotal = $0

Issue Date FY: 2025 ( Subtotal = $221,250 )
2025	2025	SLOAN-KETTERING INSTITUTE FOR CANCER RESEARCH	1275 YORK AVE	NEW YORK	NY	10065	NEW YORK	USA	Biomedical Research and Research Training	000	2	12/12/2024	NON-COMPETING CONTINUATION	$199,125
2025	2025	SLOAN-KETTERING INSTITUTE FOR CANCER RESEARCH	1275 YORK AVE	NEW YORK	NY	10065	NEW YORK	USA	Biomedical Research and Research Training	001	2	8/12/2025	NON-COMPETING CONTINUATION	$22,125
														Subtotal = $221,250

Issue Date FY: 2024 ( Subtotal = $265,500 )
2024	2024	SLOAN-KETTERING INSTITUTE FOR CANCER RESEARCH	1275 YORK AVENUE	NEW YORK	NY	10065	NEW YORK	USA	Biomedical Research and Research Training	000	1	12/27/2023	NEW	$265,500
														Subtotal = $265,500

Grand Total All Awards = $486,750

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Recombinase polymerase amplification to enable same-cell, DNA/RNA multi-omic single cell sequencing

Award Number: R21GM152950

ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

PERIOD OF PERFORMANCE START DATE: 01/01/2024

PERIOD OF PERFORMANCE END DATE: 06/30/2026

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