Dataset Information

Quantifying orthogonal barcodes for sequence census assays.

ABSTRACT:

Summary

Barcode-based sequence census assays utilize custom or random oligonucloetide sequences to label various biological features, such as cell-surface proteins or CRISPR perturbations. These assays all rely on barcode quantification, a task that is complicated by barcode design and technical noise. We introduce a modular approach to quantifying barcodes that achieves speed and memory improvements over existing tools. We also introduce a set of quality control metrics, and accompanying tool, for validating barcode designs.

Availability and implementation

https://github.com/pachterlab/kb_python, https://github.com/pachterlab/qcbc.

SUBMITTER: Booeshaghi AS

PROVIDER: S-EPMC10783946 | biostudies-literature | 2024

REPOSITORIES: biostudies-literature

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Publications

Quantifying orthogonal barcodes for sequence census assays.

Booeshaghi A Sina AS Min Kyung Hoi Joseph KHJ Gehring Jase J Pachter Lior L

Bioinformatics advances 20231220 1

<h4>Summary</h4>Barcode-based sequence census assays utilize custom or random oligonucloetide sequences to label various biological features, such as cell-surface proteins or CRISPR perturbations. These assays all rely on barcode quantification, a task that is complicated by barcode design and technical noise. We introduce a modular approach to quantifying barcodes that achieves speed and memory improvements over existing tools. We also introduce a set of quality control metrics, and accompanyin ...[more]

PMID: 38213823

Similar Datasets

Project description:BackgroundMassively parallel reporter assays (MPRAs) are an experimental technology for measuring the activity of thousands of candidate regulatory sequences or their variants in parallel, where the activity of individual sequences is measured from pools of sequence-tagged reporter genes. Activity is derived from the ratio of transcribed RNA to input DNA counts of associated tag sequences in each reporter construct, so-called barcodes. Recently, tools specifically designed to analyze MPRA data were developed that attempt to model the count data, accounting for its inherent variation. Of these tools, MPRAnalyze and mpralm are most widely used. MPRAnalyze models barcode counts to estimate the transcription rate of each sequence. While it has increased statistical power and robustness against outliers compared to mpralm, it is slow and has a high false discovery rate. Mpralm, a tool built on the R package Limma, estimates log fold-changes between different sequences. As opposed to MPRAnalyze, it is fast and has a low false discovery rate but is susceptible to outliers and has less statistical power.ResultsWe propose BCalm, an MPRA analysis framework aimed at addressing the limitations of the existing tools. BCalm is an adaptation of mpralm, but models individual barcode counts instead of aggregating counts per sequence. Leaving out the aggregation step increases statistical power and improves robustness to outliers, while being fast and precise. We show the improved performance over existing methods on both simulated MPRA data and a lentiviral MPRA library of 166,508 target sequences, including 82,258 allelic variants. Further, BCalm adds functionality beyond the existing mpralm package, such as preparing count input files from MPRAsnakeflow, as well as an option to test for sequences with enhancing or repressing activity. Its built-in plotting functionalities allow for easy interpretation of the results.ConclusionsWith BCalm, we provide a new tool for analyzing MPRA data which is robust and accurate on real MPRA datasets. The package is available at https://github.com/kircherlab/BCalm .

Dataset Information

Quantifying orthogonal barcodes for sequence census assays.

Summary

Availability and implementation

Publications

Quantifying orthogonal barcodes for sequence census assays.

Similar Datasets

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