Next-Generation DNA Barcoding for Fish Identification Using High-Throughput Sequencing in Tai Lake, China

Abstract

Tai Lake, an important biodiversity hotspot of the lower reaches of the Yangtze River in China, possesses its characteristic fish fauna. Barcoding on native species is important for species identification and biodiversity assessment with molecular-based methods, such as environmental DNA (eDNA) metabarcoding. Here, DNA-barcoding coupled with high-throughput sequencing (HTS) and traditional Sanger sequencing was introduced to barcoding 180 specimens belonging to 33 prior morphological species, including the most majority of fish fauna in Tai Lake. HTS technology, on the one hand, significantly enhances the capture of barcode sequences of fish. The successful rate of fish barcoding was 74% and 91% in Sanger and HTS, respectively. On the other hand, the HTS output has a large number (64%) of insertions and deletions, which require strict bioinformatics processing to ensure that the ‘‘true’’ barcode sequence is captured. Cross-contamination and parasites were the primary error sources that compromised attempts at the DNA barcoding of fish species. The barcode gap analysis was 100% successful at delimiting species in all specimens. The automatic barcode gap discovery (ABGD) method grouped barcode sequences into 34 OTUs, and some deep divergence and closed species failed to obtain corresponding OTUs. Overall, the local species barcode library established by HTS barcoding here is anticipated to shed new light on conserving fish diversity in Tai Lake.