Rapid mitochondrial genome sequencing based on Oxford nanopore sequencing and a proxy for vertebrate species identification using MinION device
Franco Sierra, Nicolás David
Magíster en Ciencias de la Tierra
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Molecular information is crucial for species identification when facing challenging morphology-based specimen identification. The use of DNA barcodes partially solves this problem, but in some cases when PCR is not an option (i.e. primers are not available, problems in reaction standardization) amplification-free approaches could lead to better and rapid results. Recent advances in DNA sequencing, like MinION device from Oxford Nanopore Technologies (ONT), allow to obtain genomic data with low laboratory and technical requirements, and at a relative low cost. In this study, we explore ONT sequencing for molecular species identification from a total DNA sample obtained from a neotropical rodent and also we test the technology for complete mitochondrial genome reconstruction via genome skimming. We were able to obtain “de novo” the complete mitogenome of Melanomys caliginosus (Cricetidae: Sigmodontinae) with an average depth coverage of 77.5X using ONT-only data and by combining multiple assembly routines to produce a coherent assembly (complete ORFs and RNAs). Our routine for an automated species identification was able to identify the sample with high certainty using unassembled sequence data in a reasonable computing time (1:56:31) and it was significatively reduced when a priori information related to the organism identity was known (00:04:57 and 00:01:47 when order and family were known, respectively). Our findings suggest ONT sequencing as a suitable candidate to solve species identification problems in metazoan non-model organisms and generate complete mtDNA datasets.