# Profiling Transcriptional Heterogeneity in Microbial Cells at Single Cell resolution and High-throughput using Droplet Microfluidics

> **NIH NIH DP2** · UNIVERSITY OF CHICAGO · 2020 · $2,430,000

## Abstract

PROJECT SUMMARY
Microbial transcriptional dynamics have been known to be highly dynamic and heterogeneous between cells,
seen from microscopy based and targeted sequencing approaches. Such heterogeneity can be an asset or a
liability; from a fitness perspective, transcriptional heterogeneity is a prerequisite for survival under changing
environments; however, the modern scourge of antibiotic resistance may be ascribed to such heterogeneities.
In either case, there is an enormous need to characterize the transcriptional dynamics at the resolution of
individual microbial cell. However traditional approaches relying on one or a few selected reporter genes are
inadequate for this challenge.
Recent technical advances now allow us to use RNA-Seq to profile single mammalian cells. Massive and early
barcoding followed by pooling or manipulation by microfluidics have increased the scale to tens of thousands
of cells. However, these technologies have thus far failed to translate to single microbial cells due to (1)
difficulty in single microbial cell lysis, especially those with thick cell wall; (2) difficulty to capture and barcode
relatively sparse microbial mRNAs, especially when lacking polyA tails (in bacteria); and (3) large population
size and complexity of microbial population that require orders of magnitude more cells be sampled in an
experiment.
We will leverage droplet microfluidics, develop physical, chemical and enzymatic lysis methods, and
investigate novel molecular biology and sequencing techniques to develop a single-cell microbial genomics
pipeline to (1) Isolate and (2) lyse single microbial cells; (3) capture and barcode the mRNA of single
microbial cells; and (4) process 104-105 cells per sample with hundreds of distinct transcripts per cell.
Barcoded RNA will then be pooled and sequenced at high depth. These tools will be modular and have broad
applicability beyond RNA-Seq, including single cell epigenomics and proteomics. New physical lysis modes
investigated will include MEMS, laser ablation, acoustic waves, and plasmon resonance.
The proposed project will significantly advance current technologies which are limited in throughput or the
number of RNA molecules measured. Currently, there is no successful strategy for single microbial cell RNA-
Seq at scale. Our strategy will enable cost-effective and generalized single-cell RNA-Seq in microbes at
massive throughput. Novel, hybrid microfluidic devices containing silicon, nanomaterials and elastomeric
components will be developed for single microbial cell lysis, barcoding and library prep.

## Key facts

- **NIH application ID:** 10002886
- **Project number:** 1DP2AI158157-01
- **Recipient organization:** UNIVERSITY OF CHICAGO
- **Principal Investigator:** Anindita Basu
- **Activity code:** DP2 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $2,430,000
- **Award type:** 1
- **Project period:** 2020-08-20 → 2025-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10002886

## Citation

> US National Institutes of Health, RePORTER application 10002886, Profiling Transcriptional Heterogeneity in Microbial Cells at Single Cell resolution and High-throughput using Droplet Microfluidics (1DP2AI158157-01). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10002886. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*