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The conversion of transient information into long-lasting responses is a common aspect of many biological processes and is crucial for the design of sophisticated synthetic circuits. Genomic DNA provides a rich medium for the storage of information in living cells. However, current cellular memory technologies are limited in their storage capacity and scalability.
We converted genomic DNA into a “tape recorder” for memorizing information in living cell populations. This was achieved via SCRIBE (Synthetic Cellular Recorders Integrating Biological Events), a programmable and modular architecture for generating single-stranded DNA (ssDNA) inside of living cells in response to gene regulatory signals. When coexpressed with a recombinase, these ssDNAs address specific target loci on the basis of sequence homology and introduce precise mutations into genomic DNA, thus converting transient cellular signals into genomically encoded memory. This distributed biological memory leverages the large number of cells in bacterial cultures and encodes information into their collective genomic DNA in the form of the fraction of cells that carry specific mutations.
We show that SCRIBE enables the recording of arbitrary transcriptional inputs into DNA storage registers in living cells by translating regulatory signals into ssDNAs. In Escherichia coli, we expressed ssDNAs from engineered retrons that use a reverse transcriptase protein to produce hybrid RNA-ssDNA molecules. These intracellularly expressed ssDNAs are targeted into specific genomic loci where they are recombined and converted into permanent memory. We show that genomically stored information can be readily reprogrammed by changing the ssDNA template and controlled via both chemical and light inputs. We demonstrate that genomically encoded memory can be read with a variety of techniques, including reporter genes, functional assays, and high-throughput DNA sequencing.
SCRIBE enables the recording of analog information such as the magnitude and time span of exposure to an input. This convenient feature is facilitated by the intermediate recombination rate of our current system (~10–4 recombination events per generation), which we validated via a mathematical model and computer simulations. For example, we stored the overall exposure time to chemical inducers in the DNA memory of bacterial populations for 12 days (~120 generations), independently of the induction pattern. The frequency of mutants in these populations was linearly related to the total exposure time.
Furthermore, we demonstrate that SCRIBE-induced mutations can be written and erased and can be used to record multiple inputs across the distributed genomic DNA of bacterial populations. Finally, we show that SCRIBE memory can be decomposed into independent “input,” “write,” and “read” operations and used to create genetic “logic-and-memory” circuits, as well as “sample-and-hold” circuits.
We describe a scalable platform that uses genomic DNA for analog, rewritable, and flexible memory distributed across living cell populations. We anticipate that SCRIBE will enable long-term cellular recorders for environmental and biomedical applications. Future optimization of recombination efficiencies achievable by SCRIBE could lead to more efficient single-cell digital memories and enhanced genome engineering technologies. Furthermore, the ability to regulate the generation of arbitrary targeted mutations with other gene-editing technologies should enable genomically encoded memory in additional organisms.
Cellular memory is crucial to many natural biological processes and sophisticated synthetic biology applications. Existing cellular memories rely on epigenetic switches or recombinases, which are limited in scalability and recording capacity. In this work, we use the DNA of living cell populations as genomic “tape recorders” for the analog and distributed recording of long-term event histories. We describe a platform for generating single-stranded DNA (ssDNA) in vivo in response to arbitrary transcriptional signals. When coexpressed with a recombinase, these intracellularly expressed ssDNAs target specific genomic DNA addresses, resulting in precise mutations that accumulate in cell populations as a function of the magnitude and duration of the inputs. This platform could enable long-term cellular recorders for environmental and biomedical applications, biological state machines, and enhanced genome engineering strategies.
Record your memories with a DNA recorder
DNA-based memory devices are not optimal for recording analog information, such as the magnitude of inputs over time. Farzadfard and Lu converted genomic DNA into a “tape recorder” within living bacterial populations (see the Perspective by Ausländer and Fussenegger). Specific DNAs were used to introduce precise mutations into genomic DNA. The stored information could be read out via reporter genes, functional assays, and DNA sequencing. This approach allowed the memorization of multiple inputs at the population level. The record could also be erased when required.