The complete mitogenome of Cylindrus obtusus (Helicidae, Ariantinae) using Illumina next generation sequencing.
The complete mitogenome of Cylindrus obtusus (Helicidae, Ariantinae) using Illumina next generation sequencing.
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DOI：
10.1186/1471-2164-13-114
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发表时间：
2012-03-26
期刊：
影响因子：
4.4
通讯作者：
Schilthuizen M
中科院分区：
文献类型：
--
作者：
Groenenberg DS;Pirovano W;Gittenberger E;Schilthuizen M
这项研究描述了如何完整的有丝分裂基因组的陆生蜗牛，钝头叉尾草（Draparnaud，1805年）的测序没有PCR收集标本，已在70%的乙醇8年。用Illumina GAIIx霰弹枪测序获得有丝分裂基因组。虽然使用的标本是最近收集的，并保存在DNA友好的防腐剂（而不是福尔马林，因为经常使用的旧博物馆标本），我们认为，排除PCR的促进NGS（下一代测序）消除了一个很大的障碍，在DNA测序的收集标本。我们的Illumina GAIIx方法与使用Roche 454-FLX平台的类似研究进行了简要比较。线粒体DNA序列分析表明，C. obtusus长14，610个碱基（比迄今报道的其他柱眼动物有丝分裂基因组大约0.5kb），包含后生动物典型的37个基因（13个蛋白质编码基因、2个rRNA和22个tRNA）。除了tRNA-Pro和tRNA-Ala的位置互换外，C. obtusus与报道的Cepaea nemoralis相同。与其他Sigmuratura（和大多数腹足类动物）相比，tRNA-Thr和COIII的“异常”重排并不是C. nemoralis（Helicinae亚科），但也显示出在C. obtusus（亚族Ariantinae），可能是螺旋科Helicidae的一个共形。自然历史收藏品可能为进化遗传学领域提供丰富的信息，但很难从这些标本中扩增DNA（例如由于DNA降解）。由于NGS技术不依赖于引物定向扩增（PCR），并允许DNA片段化（DNA在文库制备过程中被剪切），NGS可能是从此类标本中检索DNA序列数据的有价值的工具。Illumina GAIIx和Roche 454平台之间的比较表明，前者可能更适合有丝分裂基因组的从头测序。
This study describes how the complete mitogenome of a terrestrial snail, Cylindrus obtusus (Draparnaud, 1805) was sequenced without PCRs from a collection specimen that had been in 70% ethanol for 8 years. The mitogenome was obtained with Illumina GAIIx shot gun sequencing. Although the used specimen was collected relatively recently and kept in a DNA-friendly preservative (not formalin as frequently used with old museum specimens), we believe that the exclusion of PCRs as facilitated by NGS (Next Generation Sequencing) removes a great obstacle in DNA sequencing of collection specimens. A brief comparison is made between our Illumina GAIIx approach and a similar study that made use of the Roche 454-FLX platform. The mtDNA sequence of C. obtusus is 14,610 bases in length (about 0.5 kb larger than other stylommatophoran mitogenomes reported hitherto) and contains the 37 genes (13 protein coding genes, two rRNAs and 22 tRNAs) typical for metazoans. Except for a swap between the position of tRNA-Pro and tRNA-Ala, the gene arrangement of C. obtusus is identical to that reported for Cepaea nemoralis. The 'aberrant' rearrangement of tRNA-Thr and COIII compared to that of other Sigmurethra (and the majority of gastropods), is not unique for C. nemoralis (subfamily Helicinae), but is also shown to occur in C. obtusus (subfamily Ariantinae) and might be a synapomorphy for the family Helicidae. Natural history collections potentially harbor a wealth of information for the field of evolutionary genetics, but it can be difficult to amplify DNA from such specimens (due to DNA degradation for instance). Because NGS techniques do not rely on primer-directed amplification (PCR) and allow DNA to be fragmented (DNA gets sheared during library preparation), NGS could be a valuable tool for retrieving DNA sequence data from such specimens. A comparison between Illumina GAIIx and the Roche 454 platform suggests that the former might be more suited for de novo sequencing of mitogenomes.
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