Post-translation digital data encoding into the genomes of mammalian cell populations

High resolution cellular signal encoding is critical for better understanding of complexbiological phenomena. DNA-based biosignal encoders alter genomic or plasmid DNA ina signal dependent manner. Current approaches involve the signal of interest affecting aDNA edit by interacting with a signal specific promoter which then results in expressionof the effector molecule (DNA altering enzyme). Here we present a proof of concept of abiosignal encoding system where the enzyme terminal deoxynucleotidyl transferase(TdT) acts as the effector molecule upon directly interacting with the signal of interest. Atemplate independent DNA polymerase (DNAp), TdT incorporates nucleotides at the 3'OH ends of DNA substrate in a signal dependent manner. By employing CRISPR-Cas9to create double stranded breaks in genomic DNA we make 3'OH ends available to act assubstrate for TdT. We show that this system can successfully resolve and encodedifferent concentrations of various biosignals into the genomic DNA of HEK-293T cells.Finally, we develop a simple encoding scheme associated with the tested biosignals andencode the message "HELLO WORLD" into the genomic DNA of HEK-293T cells at apopulation level with 91% accuracy. This work demonstrates a simple and engineerablesystem that can reliably store local biosignal information into the genomes of mammaliancell populations.