High Accuracy Nanopore Sequencing.
高精度纳米孔测序。
基本信息
- 批准号:10457928
- 负责人:
- 金额:$ 88.16万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2009
- 资助国家:美国
- 起止时间:2009-09-23 至 2025-05-31
- 项目状态:未结题
- 来源:
- 关键词:AcademiaAlgorithmsBase SequenceComplexDNADNA sequencingDependenceDetectionDevelopmentEngineeringEnzyme KineticsEnzymesEpigenetic ProcessFundingGene DuplicationGenomic DNAGoalsHealthcareHybridsIndustrializationIndustryIonsKineticsLaboratoriesLengthMapsMeasuresMethodsModalityModificationMolecular MotorsMotionMotivationMotorMovementMutationMycobacterium smegmatisNational Human Genome Research InstituteNucleic AcidsNucleotidesPlayPoint MutationPolymerasePrincipal InvestigatorProbabilityProteinsRNARNA SplicingReaderResearchResearch PersonnelResolutionRoleRotationRunningSamplingSideSignal TransductionSpecificitySystemTechniquesTechnologyTestingTimeVDAC1 geneVertebral columnWalkingWorkbaseclinical applicationconstrictioncostdesignflexibilityhelicaseimprovedinnovationmarkov modelmotor behaviormutantnanoporeneural networknovelnucleobasephysical separationportabilitypreventprogramssequencing platformsingle moleculesuccesstoolvoltage
项目摘要
PROJECT SUMMARY
The objective of this proposal is to unlock the full potential of nanopore sequencing.
This sequencing method has transformative intrinsic qualities, such as long read length,
direct epigenetic modification detection, fast sample-to-answer, portability and low cost-
to-entry, but relatively low sequencing accuracy remains a significant drawback.
With NHGRI funding, we have built a team that has played a pivotal role in
developing nanopore sequencing, first engineering the highly sensitive pore MspA to
provide single-nucleotide resolution and then combining it with enzyme control for the
first proof-of-concept of nanopore sequencing. Recently, we developed a hybrid-
voltage-enzyme control method that provided a substantial increase in nanopore
sequencing accuracy. Concurrently, we developed a nanopore single-molecule tool that
measures the kinetics of enzymes which move along DNA or RNA at unprecedented
detail.
Here we build on our previous research and propose well-founded and innovative
methods to further increase the accuracy of nanopore sequencing with the ultimate goal
of a single-passage base calling accuracy of around 99%.
Our specific aims are (1) to use sequence-dependent enzyme kinetics to create a
second reader in the nanopore system that runs in tandem to the ion current reader.
This reader will provide sequence information that is independent of the ion current
reader. We will engineer enzymes with accentuated sequence-dependent kinetics. (2)
We will systematically map the sequence-dependent enzyme kinetics and splice this
information into novel base calling algorithms. In addition, we will measure the ion
current of all four different orientations of the pore-enzyme-DNA complex to maximize
the accuracy with this new method. (3) We will engineer better sequence specificity into
the ion current reader by creating a robust platform that permits asymmetric assembly
of MspA. We will then design MspA with an asymmetric pore constriction to reduce
Brownian motion of the DNA and enhance nucleotide recognition.
Our team's success to date has enabled us to form partnerships with prominent
collaborators and to gain support from many excellent labs in academia and industry,
whose expertise assists us in each step forward. We will work with our partner labs to
complete the aims outlined in this proposal.
项目总结
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(15)
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JENS GUNDLACH其他文献
JENS GUNDLACH的其他文献
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