#CONSENSUS SEQUENCE SOFTWARE#

While ONTвЂ™s MinION still shows higher error rates >5% (Wick etВ al., 2018), the new PacBio HiFi mode allows for the generation of read with <1% error (Wenger etВ al., 2019), which will greatly improve the generation of accurate DNA barcodes. These platforms offer the advantage of longer reads, at the cost of sequencing errors. Currently, two platforms are readily available for DNA barcoding efforts, PacBio's Sequel II and ONTвЂ™s MinION. Third-generation sequencing is able to sequence millions of single molecules up to several Mbs in lengths (Jain etВ al., 2018). Besides the documentation of biodiversity, this method and other amplicon sequencing technologies have been widely used for monitoring of invasive species, detection of pathogens in environmental samples, and many other applications in taxonomy, medicine, or evolutionary biology (e.g., reviewed in Kress etВ al., 2015). This technique utilizes amplification of standardized genetic regions to characterize species present within biological samples. The development of DNA barcoding (Hebert etВ al., 2003) was a major step toward large-scale characterizations of biodiversity. The identification of species in our environments is paramount to informing conservation policy and practice. We are in the middle of a biodiversity crisis, in which anthropogenic change is driving many species to extinction, often faster than they can be characterized (see, e.g., Ceballos etВ al., 2020).

#CONSENSUS SEQUENCE SOFTWARE#

We show that NGSpeciesID produces consensus sequences with improved usability by minimizing preprocessing and software installation and scalability by enabling rapid processing of hundreds to thousands of samples, while maintaining similar consensus accuracy as current pipelines. The tool includes clustering of the reads to help filter out contaminants or reads with high error rates and employs polishing strategies specific to the appropriate sequencing platform. Here, we present NGSpeciesID, a program which can generate highly accurate consensus sequences from long-read amplicon sequencing technologies, including ONT and PacBio. However, the relatively high error rates associated with these technologies still pose challenges for generating high-quality consensus sequences. This is not only advantageous for genome sequencing projects, but also advantageous for amplicon-based high-throughput sequencing experiments, such as DNA barcoding.

These platforms can generate millions of long-read sequences. Third-generation sequencing technologies, such as Oxford Nanopore Technologies (ONT) and Pacific Biosciences (PacBio), have gained popularity over the last years.

Consensus sequence