Chromatin structure analysis enables detection of DNA insertions into the mammalian nuclear genome

Abstract

BackgroundGenetically modified organisms (GMOs) have numerous biomedical, agricultural and environmental applications. Development of accurate methods for the detection of GMOs is a prerequisite for the identification and control of authorized and unauthorized release of these engineered organisms into the environment and into the food chain. Current detection methods are unable to detect uncharacterized GMOs, since either the DNA sequence of the transgene or the amino acid sequence of the protein must be known for DNA-based or immunological-based detection, respectively.MethodsHere we describe the application of an epigenetics-based approach for the detection of mammalian GMOs via analysis of chromatin structural changes occurring in the host nucleus upon the insertion of foreign or endogenous DNA.ResultsImmunological methods combined with DNA next generation sequencing enabled direct interrogation of chromatin structure and identification of insertions of various size foreign (human or viral) DNA sequences, DNA sequences often used as genome modification tools (e.g. viral sequences, transposon elements), or endogenous DNA sequences into the nuclear genome of a model animal organism.ConclusionsThe results provide a proof-of-concept that epigenetic approaches can be used to detect the insertion of endogenous and exogenous sequences into the genome of higher organisms where the method of genetic modification, the sequence of inserted DNA, and the exact genomic insertion site(s) are unknown.General significanceMeasurement of chromatin dynamics as a sensor for detection of genomic manipulation and, more broadly, organism exposure to environmental or other factors affecting the epigenomic landscape are discussed.

Advances in identifying GM plants: toward the routine detection of ‘hidden’ and ‘new’ GMOs

“…More generally, any sequence insertion (endogenous or heterologous, and therefore trans-, cis-or intragenesis) in the genome is accompanied by genetic and epigenetic unintentional modifications, sometimes distal, which are identifiable patterns even for unknown GMOs (Ashapkin et al, 2016;Filipecki and Malepszy, 2006;Kosicki et al, 2018;Ledford, 2020;Sullivan et al, 2015). This is in line with the self/non-self DNA and RNA recognitions (Monticolo et al, 2020).…”

Agnostic detection of genomic alterations by holistic DNA structural interrogation

“…TADs are organized into larger, functionally related compartments [10, 8]. As all of these levels of organization are affected by the underlying DNA sequence, measurable variations in these structures may provide new power in mutation detection as well as allow for the characterization of the effect of environmental exposure on the epigenomic landscape, as we previously suggested [11].…”

“…Genomic and epigenomic perturbations can result from exposure to a variety of environmental factors [12–14], physiological stressors [15–17] as well as direct manipulation of the host organism (e.g., genome editing) [11, 18–20]. Genome editing mediated by CRISPR/Cas9 has enabled the manipulation of nearly any organism and has been widely utilized in basic research through the modification of cell lines and model organisms [18].…”