人工合成基因组

人工生命 artificial lifen 2007年 6月 美国科学家克雷格 文特尔和他领导的研究小组宣布，他们首次实现了完整的基因组在物种间的移植，这一成功为首个 人造生命 的降生奏响了序曲。 文特尔说，这次成功让他向着制造出首个 人造生命 又迈进一步，他将在几个月内利用人工合成的基因组展开类似的移植试验，实现科研史上零的突破。如果试验成功，文特尔就能宣布他造出全球第一个 合成生命 形式。n 2010年 5月 20日 J. Craig Venter Institute (JCVI), a not-for-profit genomic research organization, published results today describing the successful construction of the first self-replicating, synthetic bacterial cell. The team synthesized the 1.08 million base pair(1080kb) chromosome of a modified Mycoplasma mycoides genome. The synthetic cell is called Mycoplasma mycoides JCVI-syn1.0 and is the proof of principle that genomes can be designed in the computer, chemically made in the laboratory and transplanted into a recipient cell to produce a new self-replicating cell controlled only by the synthetic genome. n This research will be published by Daniel Gibson et al in the May 20th edition of Science Express and will appear in an upcoming print issue of Science. Genome transplantation in bacteria: changing one species to another天然完整基因组种间转移 :As a step toward propagation of synthetic genomes, we completely replaced the genome of a bacterial cell with one from another species by transplanting a whole genome as naked DNA. Intact genomic DNA from Mycoplasma mycoides（ 蕈状支原体 ） large colony (LC), virtually free of protein, was transplanted into Mycoplasma capricolum（ 山羊支原体） cells by polyethylene glycol(PEG)-mediated transformation. Cells selected for tetracycline resistance, carried by the M. mycoides LC chromosome, contain the complete donor genome and are free of detectable recipient genomic sequences. These cells that result from genome transplantation are phenotypically identical to the M. mycoides LC donor strain as judged by several criteria.- Science. 2007 Aug 3; 317:632-8合成基因组 -Science论文Science 2 July 2010: Vol. 329. no. 5987, pp. 52 - 56 Creation of a Bacterial Cell Controlled by a Chemically Synthesized GenomeDaniel G. Gibson,1 John I. Glass et alWe report the design, synthesis, and assembly of the 1.08megabase pair Mycoplasma mycoides JCVI-syn1.0 genome starting from digitized genome sequence information and its transplantation into a M. capricolum recipient cell to create new M. mycoides cells that are controlled only by the synthetic chromosome. The only DNA in the cells is the designed synthetic DNA sequence, including “watermark“ sequences and other designed gene deletions and polymorphisms, and mutations acquired during the building process. The new cells have expected phenotypic properties and are capable of continuous self-replication. 合成生命的基本操作程序酵母中克隆细菌全基因组策略Nucleic Acids Research, 2010, Vol. 38, No. 8酵母中克隆支原体全基因组Nucleic Acids Research, 2010, Vol. 38, No. 8合成生命的关键步骤根据解读的蕈状支原体（ M. mycoides）基因组 DNA顺序，文特尔团队设计了 1078个 DNA卡盒（ cassette） ,每个 DNA卡盒长 1080bp，其末端有 80bp的顺序重叠 .这些 DNA卡盒由 JCVI指定的公司 lue Heron Biotechnology合成。随后进行三个操作步骤：第一步，从 1078个 DNA卡盒中每次取 10个构建了 109个长 10kb的 DNA卡盒 . 第二步，从 110个 10kbDNA卡盒中每次取 10个构建了 11个长 100kb的 DNA卡盒 .第三步，将 11个长 100kb的 DNA卡盒在酵母细胞中彼此连接装配成完整的人工基因组 ,全长 1080kb,以酵母人工染色体（ YAC）的形式扩增。 在酵母细胞中人工基因组 DNA不发生甲基化，当进入受体细菌后将被受体限制酶降解 ,因此需要体外甲基化加以保护。酵母人工染色体（ YAC）可以随同染色体复制扩增。因带有标记基因，可在筛选培养基上稳定遗传。从酵母细胞中分离扩增的人工合成蕈状支原体基因组 DNA，随后将人工合成基因组导入山羊支原体（ Mycoplasma capricolum）受体细胞， 山羊支原体受体细胞中编码限制性核酸内切酶基因已失活 。人工合成的蕈状支原体基因组 DNA在山羊支原体受体细胞中可以转录 mRNA,并可翻译成蛋白质。转化的山羊支原体受体细胞基因组 DNA或者被蕈状支原体限制性核酸内切酶破坏，或者在细胞繁殖后丟失。在筛选培养基上，最后只生长人工合成蕈状支原体基因组的细胞克隆。最初的实验中，人工合成的基因组并没有产上预想的结果，没有细胞生长。为此文特尔团队设计了一种检测合成的每个 DNA卡盒中是否存在错误的方法。他们将天然的 11个 100kb的 DNA卡盒和人工合成的 11个 100 kb的卡盒进行搭配重组，用于检测人工合成的 100kb卡盒中是否存在错误. 凡是检测到功能有缺陷的人工卡盒，随即进行 DNA测序，找出存在的突变并给予矫正 , 确保进入人工基因组合成的卡盒完全正确。First Self-Replicating Synthetic Bacterial Cell 20-May-2010 合成生命操作图解创造这个可复制的试验性单细胞生物花费了 4,000万美元 取名 Synthia:人造儿 合成基因组结构图人工合成基因组中的 水印The secret amino acid messages contained in “watermarks“ that were embedded in the worlds first manmade bacterial genome.NCBI checked into the genetic sequence submitted by Venters Institute and found the watermarks hidden in plain sight. The five code