Reiterative short read sequencing inside a cellular sample

Abstract

The present disclosure provides methods for conducting in situ multiplex and multi-omics detection and identification using coded padlocks probes. The methods comprise simultaneous use of RNA-specific padlock probes and polypeptide-specific padlock probes to detect both RNA and polypeptides in a cellular sample. Both types of probes include a barcode that unique identifies the RNA or polypeptide that that padlock probe detects. Both types of probes also include a batch-specific sequencing primer binding site to enable sequencing a desired subset of concatemer template molecules. Use of the batch-specific sequencing primers reduces overcrowding signals and images, to produces optical images that are intense and resolvable. By conducting multiple rounds of sequencing on the same cellular sample using different batch-specific sequencing primers enables multiplex and multi-omics sequencing to reveal numerous target RNAs and their encoded polypeptides.

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