Why not end the day with The Turtle Project! Tonight I watched a session from the Oxford Nanopore Technologies (ONT) channel titled “The Turtle Project: Conservation epigenomics of endangered sea turtles with temperature-dependent sex determination.” Eugenie ‘Charley’ Yen from the Queen Mary University of London was the presenter. They started with a slide animation of climate change and explaining temperature-dependent sex determination (TSD). With global warming, skewed population sex ratios are a major concern. Sea turtles are classified as “vulnerable to critically endangered.” Their TSD system produces males at cooler temperatures and females at higher temperatures. In the Great Barrier Reef, green sea turtles were reported to be “>99% of juveniles and sub-adults are female.” But Yen explained that the mathematical modeling used to determine these values may be misleading. Currently there is no way to determine sex that is not lethal or involves tricky interventions. Yen and team think methylation of DNA can be used to determine sex. The team studied loggerhead sea turtles of Cabo Verde. They began by creating a high-quality reference by extracting blood from a nesting female. They extracted high-molecular weight DNA and used a PromethION 24 and long-reads (23X) for assembly with Flye and polishing with Medaka. They then used Pilon for short read polishing (74X), BlobTools for decontamination, purge_dupe for haplodization, and RagTag for reference-guided scaffolding! The assembly produced 2.15 Gbps with 98% scaffolded and 726 contigs and an N50 of 129.73 Mb. With collaborators from Florida Atlantic University, they obtained eggs from nesting females and incubated at different temperatures. They then obtained samples, performed sexing, and analyzed differentially methylated sequences. These results identified 777 sex-specific differentially-methylated sequences distributed across all chromosomes. After determining a set of sex biomarkers the team performed field incubation of clutches at different depths to obtain different temperatures. To deploy sex biomarkers for large-scale trials, they used the PromethION P2 solo. They used the Rapid Barcoding Kit 24 V14 and performed sequencing in a hotel room. The Turtle Project is developing molecular tools for sea turtle conservation, using the P2 Solo!
