Optimization and Evaluation of the bestRAD Sequencing Approach: Towards Ascertainment of the Invasion Routes of the Oriental Fruit Fly, <i>Bactrocera dorsalis</i>

Data Availability Statement: The optimised molecular laboratory protocol for the bestRAD approach (Ali et al. 2016) was deposited on the Protocols.io platform (Benoit et al. 2024) and is available at dx.doi.org/10.17504/protocols.io.rm7vz3ok4gx1/v1. Demultiplexed Illumina reads for each individual sample were deposited in the Sequence Read Archive (SRA) archive and are available on the National Center for Biotechnology Information (NCBI) server under BioProject PRJNA1085726. Our study complies with the Nagoya Protocol for the Convention on Biological Diversity, with 21 legal agreements signed between CIRAD and the government agencies from the countries providing genetic samples.

International audience

anglais. The bestRAD technique is a reduced genome representation approach with high-capacity sample multiplexing and physical isolation of biotin-labelled target DNA fragments using streptavidin beads, which should reduce total cost and genotyping errors. While we here formalise the relevance of this approach within the HTS landscape, our foremost aim was to improve its replicability, validity, and transparency. We first optimised the molecular laboratory protocol and shared the associated protocols (e.g., final detailed methodologies, quality control, best practices) under the FAIR principles. Using 84 worldwide individual samples of the Oriental fruit fly, Bactrocera dorsalis, a major invasive pest, we revealed a low rate of PCR duplicates, robustness to DNA quality and quantity, high genotype call rate, insignificant genotyping error rate, high nuclear and mitochondrial genome representativeness, and a high level of genetic information. This in-depth data quality assessment, along with total cost and handling time reduced by an estimated one-third relative to the parent RAD-Seq version, demonstrates that bestRAD is an excellent compromise between cost and quality. While we generated high-quality genomic resources for B. dorsalis, we also share details and recommendations for the bestRAD technique that can be readily used in any laboratory and applied to all organisms, even without published genome sequence.