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随着DNA测序技术的发展,生物信息学、分子遗传学、精准医学以及基因组学等新兴学科也随之涌现,推动着生命科学研究的不断深入。本文就DNA测序技术发展进程中的几代测序技术的原理及优缺点进行介绍,并对以后测序技术的发展进行展望。 相似文献
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杂交测序——DNA测序新策略 总被引:2,自引:0,他引:2
杂交测序──DNA测序新策略陈尚武,马涧泉(中山医科大学生化教研室,广州510089)关键词DNA,测序,杂交人类基因组计划要求改进DNA测序方法,促进了测序技术的发展,一种全新的测序方法──杂交测序(sequencingbyhybridizatio... 相似文献
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DNA测序技术是现代生命科学研究的重要技术之一。本文对Derwent数据库中收录的,与DNA测序技术领域相关的专利申请数据进行了研究,分别从专利申请量及年度变化、生命周期、专利权人和专利发明人、专利的国际专利分类及德温特分类号等角度深入分析了DNA测序技术专利的整体产出情况、重点技术领域和主要申请机构的专利战略布局情况。通过研究发现DNA测序技术近年来发展迅速,但是主要推动者是经济发达国家。 相似文献
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基因是人类的遗传信息的载体,其遗传和表达影响人类的繁衍及各种生命活动;个体基因组DNA序列突变往往会导致疾病的发生,获取个体的基因组DNA序列将有助于疾病的诊断.DNA测序技术潜在的医学应用前景使其近几年有了飞速的发展.本文将介绍各代DNA测序技术已经取得的进展,目前尚待克服的挑战及可能的解决办法,并着重分析最新的单分子测序技术的发展潜能及临床应用前景. 相似文献
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傅俊英赵蕴华 《现代生物医学进展》2012,12(5):958-963
DNA测序技术是遗传工程的核心技术之一,发展快速和低成本的基因测序技术成为研究焦点。美国、欧盟等发达国家和地区大力支持DNA测序技术的创新研究,并投入了大量的科研经费。在美国,国家卫生研究院(NIH)下属的国家人类基因组研究院(NHGRI)、美国能源部(DOE)以及美国科学基金委(NSF)等机构是进行DNA测序技术相关项目经费分配的主要政府部门。DNA测序作为生命科学研究的关键技术也是欧盟框架计划资助的重要内容之一,其以多个欧洲国家间合作以及产学研合作的形式开展。中国在DNA测序技术领域也开展了一些研究。 相似文献
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东方田鼠特异DNA片段的克隆及核苷酸序列分析 总被引:11,自引:1,他引:11
目的获得东方田鼠的特异DNA序列.方法Aβ基因使用PCR,基因克隆,斑点杂交,DNA序列分析,生物信息学技术.结果根据小鼠MHCⅡ外显子2及其两侧序列,合成引物并扩增东方田鼠基因组DNA,将PCR产物回收、测序后,分别设计内引物扩增东方田鼠基因组DNA,其中一对引物可得到特异性扩增带,将得到的DNA片段插入PGEM-Teasy载体,进行序列分析.用这对引物扩增人、昆明小鼠、BALB/c小鼠及C57BL/6J小鼠基因组DNA,均无扩增产物.以东方田鼠特异性扩增产物为探针进行斑点杂交,除东方田鼠基因组DNA外,其他几种动物基因组DNA均为阴性结果.进一步对该DNA片段进行了BLAST同源性搜索和外显子预测,在Genbank中没有发现高度同源序列,并且找到一个可能的外显子,该外显子由69个氨基酸组成.结论获得的DNA片段为东方田鼠的特异片段,这将为从分子水平深入研究东方田鼠的遗传背景、生物进化规律以及东方田鼠抗日本血吸虫的机理奠定基础. 相似文献
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HRP-HBVDNA探针在临检应用中的研究 总被引:2,自引:0,他引:2
本文介绍了一种简便的检测血清HBVDNA的方法。参照Renz等人的标记方法,构建了直接酶标HRP HBVDNA探针。此探针经与固定在硝酸纤维素滤膜上的血清靶DNA杂交后,可通过化学发光自显影检测技术观察结果。敏感度可检测0-1pg靶DNA,相当于同位素探针的灵敏度。对63份HBsAgHBeAg和Anti HBcELISA阳性血清以及24份HBsAgAnti HBc阳性,HbeAg阴性血清用HRP HBVDNA探针进行检测,结果探针HBVDNA阳性率分别为100%(63)和58%(14);对50份HBsAg,ELISA阴性和ALT正常的血清,探针HBVDNA全部阴性。实验结果表明本方法具有很大的推广应用价值。 相似文献
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Wei L Jiang P Xu W Li H Zhang H Yan L Chan-Park MB Liu XW Tang K Mu Y Pervushin K 《The Journal of biological chemistry》2011,286(8):6291-6300
Abnormal aggregation of islet amyloid polypeptide (IAPP) into amyloid fibrils is a hallmark of type 2 diabetes. In this study, we investigated the initial oligomerization and subsequent addition of monomers to growing aggregates of human IAPP at the residue-specific level using NMR, atomic force microscopy, mass spectroscopy, and computational simulations. We found that in solution IAPPs rapidly associate into transient low-order oligomers such as dimers and trimers via interactions between histidine 18 and tyrosine 37. This initial event is proceeded by slow aggregation into higher-order spherical oligomers and elongated fibrils. In these two morphologically distinct types of aggregates IAPPs adopt structures with markedly different residual flexibility. Here we show that the anti-amyloidogenic compound resveratrol inhibits oligomerization and amyloid formation via binding to histidine 18, supporting the finding that this residue is crucial for on-pathway oligomer formation. 相似文献
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为了分析比较食虫目高重复顺序的结构特征,我们将食虫目动物刺猬的高重复顺序DNA进行了分离、纯化、分子克隆,并测出了1186bp的核苷酸顺序。经分析发现在碱基组成上它与食虫目另一动物臭鼩有许多相似之处,如:G-C碱基对与A-T碱基对在整个片段中分布很不均衡;片段内部含有许多连续多次重复的短片段等,很可能为食虫目动物所特有。 相似文献
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A theoretical analysis was developed to predict molecular hybridization rates for microarrays where samples flow through microfluidic channels and for conventional microarrays where samples remain stationary during hybridization. The theory was validated by using a multiplexed microfluidic microarray where eight samples were hybridized simultaneously against eight probes using 60-mer DNA strands. Mass transfer coefficients ranged over three orders of magnitude where either kinetic reaction rates or molecular diffusion rates controlled overall hybridization rates. Probes were printed using microfluidic channels and also conventional spotting techniques. Consistent with the theoretical model, the microfluidic microarray demonstrated the ability to print DNA probes in less than 1 min and to detect 10-pM target concentrations with hybridization times in less than 5 min. 相似文献
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Kristina Warton Vita Lin Tina Navin Nicola J Armstrong Warren Kaplan Kevin Ying Brian Gloss Helena Mangs Shalima S Nair Neville F Hacker Robert L Sutherland Susan J Clark Goli Samimi 《BMC genomics》2014,15(1)
Background
Free circulating DNA (fcDNA) has many potential clinical applications, due to the non-invasive way in which it is collected. However, because of the low concentration of fcDNA in blood, genome-wide analysis carries many technical challenges that must be overcome before fcDNA studies can reach their full potential. There are currently no definitive standards for fcDNA collection, processing and whole-genome sequencing. We report novel detailed methodology for the capture of high-quality methylated fcDNA, library preparation and downstream genome-wide Next-Generation Sequencing. We also describe the effects of sample storage, processing and scaling on fcDNA recovery and quality.Results
Use of serum versus plasma, and storage of blood prior to separation resulted in genomic DNA contamination, likely due to leukocyte lysis. Methylated fcDNA fragments were isolated from 5 donors using a methyl-binding protein-based protocol and appear as a discrete band of ~180 bases. This discrete band allows minimal sample loss at the size restriction step in library preparation for Next-Generation Sequencing, allowing for high-quality sequencing from minimal amounts of fcDNA. Following sequencing, we obtained 37×106-86×106 unique mappable reads, representing more than 50% of total mappable reads. The methylation status of 9 genomic regions as determined by DNA capture and sequencing was independently validated by clonal bisulphite sequencing.Conclusions
Our optimized methods provide high-quality methylated fcDNA suitable for whole-genome sequencing, and allow good library complexity and accurate sequencing, despite using less than half of the recommended minimum input DNA.Electronic supplementary material
The online version of this article (doi:10.1186/1471-2164-15-476) contains supplementary material, which is available to authorized users. 相似文献20.
We describe a new DNA sequencing method called sequencing by denaturation (SBD). A Sanger dideoxy sequencing reaction is performed on the templates on a solid surface to generate a ladder of DNA fragments randomly terminated by fluorescently labeled dideoxyribonucleotides. The labeled DNA fragments are sequentially denatured from the templates and the process is monitored by measuring the change in fluorescence intensities from the surface. By analyzing the denaturation profiles, the base sequence of the template can be determined. Using thermodynamic principles, we simulated the denaturation profiles of a series of oligonucleotides ranging from 12 to 32 bases and developed a base-calling algorithm to decode the sequences. These simulations demonstrate that DNA molecules up to 20 bases can be sequenced by SBD. Experimental measurements of the melting profiles of DNA fragments in solution confirm that DNA sequences can be determined by SBD. The potential limitations and advantages of SBD are discussed. With SBD, millions of sequencing reactions can be performed on a small area on a surface in parallel with a very small amount of sequencing reagents. Therefore, DNA sequencing by SBD could potentially result in a significant increase in speed and reduction in cost in large-scale genome resequencing. 相似文献