Squeezing water from a stone: high-throughput sequencing from a 145-year old holotype resolves (barely) a cryptic species problem in flying lizards

PeerJ

article

Jimmy A. McGuire¹  Darko D. Cotoras³  Brendan O’Connell⁶  Shobi Z.S. Lawalata¹  Cynthia Y. Wang-Claypool¹  Alexander Stubbs¹  Xiaoting Huang⁹  Guinevere O.U. Wogan^1,10  Sarah M. Hykin¹  Sean B. Reilly¹  Ke Bi^1,11  Awal Riyanto^1,12  Evy Arida^1,12  Lydia L. Smith¹  Heather Milne⁶  Jeffrey W. Streicher^1,13  Djoko T. Iskandar^1,14 

[1] Museum of Vertebrate Zoology, University of California;Department of Integrative Biology, University of California;Department of Ecology and Evolutionary Biology, University of California;Entomology Department, California Academy of Sciences;Center for Comparative Genomics, California Academy of Sciences;Department of Biomolecular Engineering and Bioinformatics, Baskin School of Engineering, University of California;Department of Medical & Molecular Genetics, School of Medicine, Oregon Health & Science University;Jalan Hayam Wuruk, United in Diversity Foundation;Key Laboratory of Marine Genetics and Breeding, Ocean University of China;Department of Environmental Science, Policy, and Management, University of California;Computational Genomics Resource Laboratory, California Institute for Quantitative Biosciences, University of California;Museum Zoologicum Bogoriense, Research Center for Biology-The Indonesian Institute of Sciences;Department of Life Sciences;School of Life Sciences and Technology, Institute of Technology Bandung

关键词: Draco;    Formalin-fixation;    Ancient DNA;    Phylogeny;    Taxonomy;   

DOI  :  10.7717/peerj.4470

学科分类：社会科学、人文和艺术（综合）

来源: Inra

【 摘 要 】

We used Massively Parallel High-Throughput Sequencing to obtain genetic data from a 145-year old holotype specimen of the flying lizard, Draco cristatellus. Obtaining genetic data from this holotype was necessary to resolve an otherwise intractable taxonomic problem involving the status of this species relative to closely related sympatric Draco species that cannot otherwise be distinguished from one another on the basis of museum specimens. Initial analyses suggested that the DNA present in the holotype sample was so degraded as to be unusable for sequencing. However, we used a specialized extraction procedure developed for highly degraded ancient DNA samples and MiSeq shotgun sequencing to obtain just enough low-coverage mitochondrial DNA (721 base pairs) to conclusively resolve the species status of the holotype as well as a second known specimen of this species. The holotype was prepared before the advent of formalin-fixation and therefore was most likely originally fixed with ethanol and never exposed to formalin. Whereas conventional wisdom suggests that formalin-fixed samples should be the most challenging for DNA sequencing, we propose that evaporation during long-term alcohol storage and consequent water-exposure may subject older ethanol-fixed museum specimens to hydrolytic damage. If so, this may pose an even greater challenge for sequencing efforts involving historical samples.

【 授权许可】

CC BY