Squeezing water from a stone: high-throughput sequencing from a 145-year old holotype resolves (barely) a cryptic species problem in flying lizards.
Squeezing water from a stone: high-throughput sequencing from a 145-year old holotype resolves (barely) a cryptic species problem in flying lizards.
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作者：
McGuire JA;Cotoras DD;O'Connell B;Lawalata SZS;Wang-Claypool CY;Stubbs A;Huang X;Wogan GOU;Hykin SM;Reilly SB;Bi K;Riyanto A;Arida E;Smith LL;Milne H;Streicher JW;Iskandar DT
我们使用大规模平行高重复序列测定法从一个145岁的飞蜥Draco cristatellus的正模标本中获得遗传数据。从这个正模中获得遗传数据是必要的，以解决一个棘手的分类学问题，涉及这个物种相对于密切相关的同域龙物种的地位，否则无法根据博物馆标本相互区分。初步分析表明，正模标本中的DNA已经降解，无法用于测序。然而，我们使用了一种专门为高度降解的古代DNA样本开发的提取程序和MiSeq鸟枪测序，以获得足够的低覆盖率线粒体DNA（721个碱基对）来最终解决正模标本的物种状态以及该物种的第二个已知标本。正模标本是在福尔马林固定法出现之前制备的，因此最初很可能是用乙醇固定的，从未接触过福尔马林。而传统的智慧认为，福尔马林固定的样本应该是最具挑战性的DNA测序，我们建议，蒸发在长期的酒精储存和随之而来的水暴露可能会受到较旧的乙醇固定的博物馆标本水解损伤。如果是这样，这可能会对涉及历史样本的测序工作提出更大的挑战。
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.
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影响因子：
--
作者：
Meyer, Matthias;Kircher, Martin
通讯作者：
Kircher, Martin
影响因子：
2.1
作者：
Arbeli, Ziv;Fuentes, Cilia L.
通讯作者：
Fuentes, Cilia L.
来自阿尔泰山脉的尼安德特人的完整基因组序列。
影响因子：
64.8
作者：
通讯作者：
--
DOI：
10.1093/bioinformatics/bts199
发表时间：
2012-06-15
期刊：
Bioinformatics (Oxford, England)
影响因子：
--
作者：
Kearse M;Moir R;Wilson A;Stones-Havas S;Cheung M;Sturrock S;Buxton S;Cooper A;Markowitz S;Duran C;Thierer T;Ashton B;Meintjes P;Drummond A
通讯作者：
Drummond A
影响因子：
7
作者：
Huson, Daniel H.;Auch, Alexander F.;Schuster, Stephan C.
通讯作者：
Schuster, Stephan C.