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uSort-M

Rapid and low-cost parsed variant library generation

uSort-M converts pooled DNA libraries into collections of isolated and verified sequences. uSort-M achieves significant (often >10x) cost savings compared to traditional gene synthesis. For most libraries, the time required to clone, sort, barcode, and sequence-verify thousands of sequences is often less than 10 days.

uSort-M workflow: pooled library to FACS sorting to barcoding to sequencing to arrayed clones

From a pooled DNA library (e.g., Twist Bioscience oligo pool), uSort-M uses multiplex assembly (e.g., Golden Gate or Gibson) to generate a plasmid library for transformation into E. coli. Single cells are isolated via FACS, barcoded by well-specific PCR, sequenced, and cherry-picked into a final arrayed library.

What will it cost?

My gene is bp in length and I want to parse variants.

uSort-M: 0-300 bp uses IDT oPools or Twist Oligo Pools; >300 bp uses 30 bp substitution library cloned into full-length gene.
Traditional: Direct gene fragment synthesis (Twist Gene Fragments, IDT eBlocks).

Fold sampling: 4× Library skew: 4× (Q90/Q10) Growth rate: 67% PCR failure: 3%
Synthesis Method (click to compare scenarios)
Recommended synthesis: Twist Oligo Pools (0-350 bp)
Total Cost Comparison
$3,090 uSort-M
vs
$24,050 Traditional
7.8× savings
Total Time
6 days
Cost per Variant
$6.18
vs $48.10 trad.
Estimated Timeline

Quick Start

# Install uSort-M
git clone https://github.com/FordyceLab/usortm.git
cd usortm
pip install -e ".[all]"

# Quick cost estimate
usortm estimate --library-size 500 --seq-length 300

# Plan a full experiment
usortm plan variants.csv --output my_project/

See the Getting Started guide for detailed installation and usage instructions.

Citation

If you use uSort-M in your research, please cite:

Olivas MB, Almhjell PJ, Shanahan JD, Fordyce PM. uSort-M: Scalable isolation of user-defined sequences from diverse pooled libraries. bioRxiv (2026).