Biology subjects

Volf, V.

Publications and source records attributed to Volf, V..

4 recordsLinked to original sources

bioRxiv2023

Cryptography in the DNA of living cells enabled by multi-site base editing

Although DNA is increasingly being adopted as a generalizable medium for information storage and transfer, reliable methods for ensuring information security remain to be addressed. In this study, we developed and validated a cryptographic encoding scheme, Genomic Sequence Encryption (GSE), to address the challenge of information confidentiality and integrity in biological substrates. GSE enables genomic information encoding that is readable only with a cryptographic key. We show that GSE can be used for cell signatures that enable the recipient of a cell line to authenticate its origin and validate if the cell line has been modified in the interim. We implement GSE through multi-site base editing and encode information through editing across >100 genomic sites in mammalian cells. We further present an enrichment step to obtain individual stem cells with more than two dozen edits across a single genome with minimal screening. This capability can be used to introduce encrypted signatures in living animals. As an encryption scheme, GSE is falsification-proof and enables secure information transfer in biological substrates.

synthetic biology↗

bioRxiv2020

Characterizing the portability of RecT-mediated oligonucleotide recombination

Bacterial genome editing methods are used to engineer strains for biotechnology and fundamental research. Homologous recombination (HR) is the most versatile method of genome editing, but traditional techniques using endogenous RecA-mediated pathways are inefficient and laborious. Phage encoded RecT proteins can improve HR over 1000-fold, but these proteins have limited portability between species. Using Escherichia coli, Lactococcus lactis, Mycobacterium smegmatis, Lactobacillus rhamnosus, and Caulobacter crescentus we investigated the hostlimited functionality of RecTs. We find that these proteins specifically recognize the 7 C-terminal amino acids of the bacterial single-stranded DNA-binding protein (SSB), and are portable between species only if compatibility with this host domain is maintained. Furthermore, in some species, we find that co-expressing otherwise incompatible RecTs with a paired bacterial SSB is sufficient to establish functionality. Finally, we demonstrate that high-efficiency HR surpasses the mutational capacity of more widely used error-prone methods for genome diversification, and can be used to identify exceptional phenotypes inaccessible through sequential nucleotide conversions.

synthetic biology↗

bioRxiv2019

Robust Neural Networks are More Interpretable for Genomics

Deep neural networks (DNNs) have been applied to a variety of regulatory genomics tasks. For interpretability, attribution methods are employed to provide importance scores for each nucleotide in a given sequence. However, even with state-of-the-art DNNs, there is no guarantee that these methods can recover interpretable, biological representations. Here we perform systematic experiments on synthetic genomic data to raise awareness of this issue. We find that deeper networks have better generalization performance, but attribution methods recover less interpretable representations. Then, we show training methods promoting robustness - including regularization, injecting random noise into the data, and adversarial training - significantly improve interpretability of DNNs, especially for smaller datasets.

genomics↗

bioRxiv2019

Enabling large-scale genome editing by reducing DNA nicking

To extend the frontier of genome editing and enable the radical redesign of mammalian genomes, we developed a set of dead-Cas9 base editor (dBE) variants that allow editing at tens of thousands of loci per cell by overcoming the cell death associated with DNA double-strand breaks (DSBs) and single-strand breaks (SSBs). We used a set of gRNAs targeting repetitive elements - ranging in target copy number from about 31 to 124,000 per cell. dBEs enabled survival after large-scale base editing, allowing targeted mutations at up to ~13,200 and ~2610 loci in 293T and human induced pluripotent stem cells (hiPSCs), respectively, three orders of magnitude greater than previously recorded. These dBEs can overcome current on-target mutation and toxicity barriers that prevent cell survival after large-scale genome engineering.\n\nOne Sentence SummaryBase editing with reduced DNA nicking allows for the simultaneous editing of >10,000 loci in human cells.

synthetic biology↗