What is the available evidence for the range of applications of genome-editing as a new tool for plant trait modification and the potential occurrence of associated off-target effects: a systematic map
Within the last decades, genome-editing techniques such as CRISPR/Cas, TALENs, Zinc-Finger Nucleases, Meganucleases, Oligonucleotide-Directed Mutagenesis and base editing have been developed enabling a precise modification of DNA sequences. Such techniques provide options for simple, time-saving and cost-effective applications compared to other breeding techniques and hence genome editing has already been promoted for a wide range of plant species. Although the application of genome-editing induces less unintended modifications (off-targets) in the genome compared to classical mutagenesis techniques, off-target effects are a prominent point of criticism as they are supposed to cause unintended effects, e.g. genomic instability or cell death. To address these aspects, this map aims to answer the following question: What is the available evidence for the range of applications of genome-editing as a new tool for plant trait modification and the potential occurrence of associated off-target effects? This primary question will be considered by two secondary questions: One aims to overview the market-oriented traits being modified by genome-editing in plants and the other explores the occurrence of off-target effects.
A literature search in nine bibliographic databases, Google Scholar, and 47 web pages of companies and governmental agencies was conducted using predefined and tested search strings in English language. Articles were screened on title/abstract and full text level for relevance based on pre-defined inclusion criteria. The relevant information of included studies were mapped using a pre-defined data extraction strategy. Besides a descriptive summary of the relevant literature, a spreadsheet containing all extracted data is provided.
Altogether, 555 relevant articles from journals, company web pages and web pages of governmental agencies were identified containing 1328 studies/applications of genome-editing in model plants and agricultural crops in the period January 1996 to May 2018. Most of the studies were conducted in China followed by the USA. Genome-editing was already applied in 68 different plants. Although most of the studies were basic research, 99 different market-oriented applications were identified in 28 different crops leading to plants with improved food and feed quality, agronomic value like growth characteristics or increased yield, tolerance to biotic and abiotic stress, herbicide tolerance or industrial benefits. 252 studies explored off-target effects. Most of the studies were conducted using CRISPR/Cas. Several studies firstly investigated whether sites in the genome show similarity to the target sequence and secondly analyzed these potential off-target sites by sequencing. In around 3% of the analyzed potential off-target sites, unintended mutations were detected. Only a few studies conducted off-target analyses using unbiased detection methods (e.g. whole genome sequencing). No off-target effects that could be correlated to the genome-editing process were identified in these studies.
The rapid adoption in plant breeding was demonstrated by a considerable number of market oriented applications (crops and traits) described in publications worldwide. Studies investigating off-target effects are very heterogeneous in their structure and design. Therefore, an in-depth assessment regarding their weight of evidence is mandatory.
New plant breeding techniques, Gene editing, Targeted genome modification, Mutagenesis, Unintended effects, Off-target mutation, Evidence map, Evidence synthesis
Plant breeding is a developing process and breeding methods have continuously evolved over time. In recent years, genome editing techniques such as clustered regularly interspaced short palindromic repeats/CRISPR associated proteins (CRISPR/Cas), transcription activator-like effector nucleases (TALENs), zinc-finger nucleases (ZFN), meganucleases (MN) and oligonucleotide-directed mutagenesis (ODM) enabled a precise modification of DNA sequences in plants. Genome editing has already been applied in a wide range of plant species due to its simplicity, time saving and cost-effective application compared to earlier breeding techniques including classical mutagenesis. Although genome editing techniques induce much less unintended modifications in the genome (off-target effects) compared to classical mutagenesis techniques, off-target effects are a prominent point of criticism as they might cause genomic instability, cytotoxicity and cell death.
The aim of this systematic map is to address the following primary question: “What is the available evidence for the application of genome editing as a new tool for plant trait modification and the potential occurrence of associated off–target effects”? The primary question will be considered by two secondary questions: One is aimed at the traits being modified by genome editing in plants and the other explores the occurrence of off-target effects. The systematic map will focus on model plants as well as on plants produced for agricultural production that were subjected to genome editing techniques. Academic and grey literature will be searched in English and German language. Inclusion/exclusion criteria were developed for the two secondary questions and will be applied on title/abstract and full text stage. Included studies will be catalogued in a searchable and open access database and study results will be summarized using descriptive statistics. Furthermore, the extracted data will serve as a preparatory step for further in-depth analysis, e.g. by a systematic review.