One bacterial cell, one complete genome.
One bacterial cell, one complete genome.
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DOI：
10.1371/journal.pone.0010314
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发表时间：
2010-04-23
期刊：
影响因子：
3.7
通讯作者：
Cheng JF
中科院分区：
文献类型：
--
作者：
Woyke T;Tighe D;Mavromatis K;Clum A;Copeland A;Schackwitz W;Lapidus A;Wu D;McCutcheon JP;McDonald BR;Moran NA;Bristow J;Cheng JF
虽然迄今为止测序的大部分成品微生物基因组来源于培养的细菌和古细菌代表，但绝大多数微生物逃避了当前的培养尝试，严重限制了从这些环境物种中恢复完整或甚至部分基因组的能力。单细胞基因组学是一种新的不依赖于培养的方法，它可以获得单个细胞的遗传物质。据我们所知，迄今为止还没有一个单细胞基因组被关闭和完成。在这里，我们报告了完整的基因组从一个未培养的单细胞的沟藻。对从绿色神枪手Draeculacephala minerva细菌组中分离的单共生体细胞进行数字PCR，使我们能够评估这种细菌是多倍体，每个细胞的基因组拷贝数约为200-900，使其成为单细胞精加工工作的最合适目标。对于单细胞鸟枪测序，分离单个Sulcia细胞，并通过多重置换扩增（MDA）扩增全基因组。基于桑格的精加工方法使我们能够关闭基因组。为了验证我们的单细胞基因组的正确性并排除MDA衍生的人工产物，我们使用宏基因组学方法从合并的细菌组中独立地进行鸟枪测序和组装Sulcia基因组，产生几乎相同的基因组。我们检测到的四个变化似乎是两个样本之间真正的生物学差异。单细胞基因组与细菌组宏基因组序列数据的比较检测到两个单核苷酸多态性（SNP），表明极低的遗传多样性内Sulcia人口。这项研究证明了单细胞基因组学在环境样本中产生完整，高质量，非复合参考基因组的能力，可用于群体遗传分析。
While the bulk of the finished microbial genomes sequenced to date are derived from cultured bacterial and archaeal representatives, the vast majority of microorganisms elude current culturing attempts, severely limiting the ability to recover complete or even partial genomes from these environmental species. Single cell genomics is a novel culture-independent approach, which enables access to the genetic material of an individual cell. No single cell genome has to our knowledge been closed and finished to date. Here we report the completed genome from an uncultured single cell of Candidatus Sulcia muelleri DMIN. Digital PCR on single symbiont cells isolated from the bacteriome of the green sharpshooter Draeculacephala minerva bacteriome allowed us to assess that this bacteria is polyploid with genome copies ranging from approximately 200–900 per cell, making it a most suitable target for single cell finishing efforts. For single cell shotgun sequencing, an individual Sulcia cell was isolated and whole genome amplified by multiple displacement amplification (MDA). Sanger-based finishing methods allowed us to close the genome. To verify the correctness of our single cell genome and exclude MDA-derived artifacts, we independently shotgun sequenced and assembled the Sulcia genome from pooled bacteriomes using a metagenomic approach, yielding a nearly identical genome. Four variations we detected appear to be genuine biological differences between the two samples. Comparison of the single cell genome with bacteriome metagenomic sequence data detected two single nucleotide polymorphisms (SNPs), indicating extremely low genetic diversity within a Sulcia population. This study demonstrates the power of single cell genomics to generate a complete, high quality, non-composite reference genome within an environmental sample, which can be used for population genetic analyzes.
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影响因子：
4.4
作者：
Raghunathan, A;Ferguson, HR;Lasken, RS
通讯作者：
Lasken, RS
DOI：
10.1073/pnas.0801980105
发表时间：
2008-06-10
影响因子：
11.1
作者：
Elkins, James G.;Podar, Mircea;Stetter, Karl O.
通讯作者：
Stetter, Karl O.
影响因子：
4.4
作者：
Moran, NA;Tran, P;Gerardo, NM
通讯作者：
Gerardo, NM
DOI：
10.1073/pnas.0906424106
发表时间：
2009-09-08
影响因子：
11.1
作者：
McCutcheon, John P.;McDonald, Bradon R.;Moran, Nancy A.
通讯作者：
Moran, Nancy A.
影响因子：
3.7
作者：
Rodrigue S;Malmstrom RR;Berlin AM;Birren BW;Henn MR;Chisholm SW
通讯作者：
Chisholm SW