A spatial genome aligner for multiplexed DNA-FISH

Multiplexed fluorescence in situ hybridization (FISH) has emerged as a powerful approach for analyzing 3D genome organization, but it is eminently challenging to derive chromosomal conformations from noisy fluorescence signals. Tracing chromatin is not straightforward as chromosomes lack conserved shapes for reference checking whether an observed fluorescence signal belongs to a chromatin fiber or not. Here we report a spatial genome aligner that parses true chromatin signal from noise by aligning signals to a DNA polymer model. We demonstrate that this spatial genome aligner can efficiently reconstruct chromosome architectures from DNA-FISH data across multiple scales and determine chromosome ploidies de novo in interphase cells. Reprocessing of previous whole-genome chromosome tracing data with this method revealed the spatial aggregation of sister chromatids in S/G2 phase cells in asynchronous mouse embryonic stem cells, and uncovered extranumerary chromosomes that remain tightly paired in post-mitotic neurons of the adult mouse cortex. Our spatial genome aligner may facilitate the adaption of multiplexed DNA-FISH by the community.