Improving overlap-finding techniques for whole genome shotgun data

一个基因组(细胞中的DNA)可以用一个叫做“碱基”的字母序列来表示。一个庞大的基因组可以由数十亿个碱基组成。化学技术使科学家一次只能读取几百个碱基。全基因组鸟枪(WGS)组装技术通过从基因组中随机选择这样的短片段，确定片段的序列，然后通过计算重新组装数百万个片段来创建整个基因组的序列草图。根据它们的序列比较，如果两个片段似乎来自基因组的同一部分，那么它们就被称为“重叠”。这个项目的目标是集中努力，使用复杂的纠错技术的组合，产生一组非常强大的重叠，以及通过确保两个片段来自基因组的同一附近来“定位”片段以验证重叠。首先，大多数基因组包含许多“重复区域”，即两个或更多个几乎完全相同的长序列拷贝。因此，实际上没有重叠的两个片段看起来可能是重叠的。第二，随机抽样技术导致许多碱基错误-碱基可能被误读或完全遗漏。这些错误，再加上重复区域通常略有不同的事实，使得很难区分虚假重叠和真正的重叠，其中一个或两个片段包含读取错误。因此，如果不采取极端谨慎的措施，很容易使用虚假的重叠，从而将基因组的遥远部分巧妙地连接在一起。与塞莱拉基因组学、贝勒医学院和基因组研究所(TIGR)合作的初步结果表明，研究人员目前的技术已经可以以更高的质量产生更多的序列。目标是改进这些技术并使其广泛使用。确定和解释遗传信息是21世纪的重大挑战之一。基因组，即细胞中的所有DNA，是多样性的分子基础和遗传信息的基石。人类、小鼠以及一些昆虫、鱼类、植物和细菌的基因组已经获得。这只是一个开始，但仅仅研究少数几个物种的基因组并不能完全理解生物过程。联邦政府每年花费约1亿美元来生成序列数据。数以百万计的基因组小片段是从基因组中提取的。第二个阶段被称为“组装”，当这些碎片像一个巨大的拼图游戏一样在电脑上重新组装时。这个谜题因两个事实而变得复杂：首先，许多拼图碎片有小错误，使它们与本应匹配的碎片不匹配；其次，许多不应该放在一起的碎片实际上非常适合在一起。这使得正确组装基因组变得极其困难。有两种方法可以减少歧义：第一，一种方法可以产生更多的碎片。然而，每个新零件的成本约为2美元，人们需要生成数百万个新零件才能对组装质量产生显著影响。调查人员使用了第二条路线。他们试图从现有的碎片中榨取尽可能多的信息。后一条路线要便宜得多，与现有技术相比，这里仍有很大的改进空间。研究人员正在使用复杂的数学方法，以极高的精度帮助辨别哪些片段可以拼接，哪些不能拼接。研究人员与几个大型测序中心合作的初步结果表明，使用他们的技术对片段进行“前处理”可以产生更多的基因组，错误更少。该项目旨在进一步扩展这些想法，并使所有研究人员都可以自由获取这些想法。这对联邦基因组(生物技术)项目的影响可能是巨大的。

Yorke A genome (the DNA in a cell) can be represented by asequence of letters called "bases." A large genome can consistof billions of bases. Chemical techniques allow scientists toread only a few hundred bases at a time. The whole genome shotgun(WGS) assembly technique creates a draft of the sequence of awhole genome by selecting such short fragments at random from thegenome, determining the sequence of the fragments, and thencomputationally re-assembling millions of these fragments. Twofragments are said to "overlap" if it is plausible that they comefrom the same part of the genome, based on a comparison of theirsequences. The goal of this project is to focus efforts onproducing an extremely robust set of overlaps, using acombination of sophisticated error-correction techniques, as wellas "localizing" fragments to validate overlaps by ensuring thatboth fragments come from the same vicinity of the genome.Several issues complicate the determination of which pairs offragments overlap. First, most genomes contain many "repeatregions," i.e., two or more almost identical copies of longstretches of sequence. Thus, two fragments that do not actuallyoverlap may look like they do. Second, the random samplingtechnique results in many base errors --- bases can be mis-reador missed entirely. These errors, combined with the fact thatrepeat regions usually differ slightly, make it very difficult todistinguish a spurious overlap from a true overlap in which oneor both fragments contain read errors. Thus, if extreme care isnot taken, it is easy to use a spurious overlap and therebymistakenly connect distant parts of the genome. Preliminaryresults in collaboration with Celera Genomics, the Baylor Collegeof Medicine, and The Institute for Genomic Research (TIGR) havedemonstrated that the investigator's current techniques canalready produce more sequence at higher quality. The goal isimprove these techniques and make them widely available. The determination and interpretation of genetic informationis one of the great challenges of the twenty-first century. Thegenome, i.e., all the DNA in a cell, is the molecular basis ofdiversity and the cornerstone of genetic information. Draftgenomes have been obtained for human, mouse, and some insects,fish, plants, and bacteria. This is a start, but a fullunderstanding of biological processes cannot be had by studyingthe genomes of only a handful of species. The federal governmentis spending about 100 million dollars per year generatingsequence data. Millions of small pieces of a genome are sampledfrom the genome. The second stage is called "assembly," whenthese pieces are re-assembled on a computer like a giant jigsawpuzzle. The puzzle is complicated by two facts: first, many ofthe puzzle pieces have small errors that make them mis-fitagainst pieces that they SHOULD fit with; and second, many piecesthat should NOT go together actually fit together quite well.This makes it extremely difficult to correctly assemble a genome.There are two ways to decrease the ambiguities: first, one couldgenerate more pieces. However, each new piece costs about $2,and one would need to generate millions of new pieces to have asignificant effect on assembly quality. The investigators use asecond route. They attempt to squeeze as much information out ofthe existing pieces as possible. The latter route issubstantially cheaper, and there is still much room forimprovement here over existing techniques. The investigators areusing sophisticated mathematics to help discern with extremeprecision those pairs of pieces that do, and those that do not,fit together. Preliminary results of the investigators -- incollaboration with several large sequencing centers -- havedemonstrated that using their techniques to "pre-process" thepieces can produce more of the genome, with fewer errors. Thisproject aims at extending these ideas further and making themfreely accessible to all investigators. The impact on the federalgenome (biotechnology) projects is potentially great.