Abstract
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.
Highlights
The advent of Massively Parallel High-Throughput Sequencing (HTS) has dramatically altered the manner in which geneticists conduct their research
Bi & McGuire (2015) demonstrated that low-coverage genomic sequences could be recovered from a 30-year old formalin-fixed museum specimen, though they were unsuccessful with a ∼100-year old specimen
The samples that failed in the Ruane & Austin (2016) experiment included subsets of both their formalin- and alcohol-fixed samples, indicating that old alcohol-preserved museum specimens are not necessarily less problematic than those initially fixed with formalin
Summary
The advent of Massively Parallel High-Throughput Sequencing (HTS) has dramatically altered the manner in which geneticists conduct their research. Ruane & Austin (2016) sequenced Ultra-Conserved Elements (UCEs) from both formalin-fixed (n = 11) and ethanol-fixed (n = 10) museum specimens, including one sample that was collected between 1878 and 1911 Both had mixed success, with the quantity of DNA recovered in the extraction stage likely playing the largest role in the performance of their sequencing efforts. The samples that failed in the Ruane & Austin (2016) experiment included subsets of both their formalin- (seven of 16) and alcohol-fixed (four of five) samples, indicating that old alcohol-preserved museum specimens are not necessarily less problematic than those initially fixed with formalin This is surprising given that contemporary tissue samples earmarked for genetic analysis are routinely stored in 95% ethanol
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
