RiboTag: A novel technique to profile cell type specific gene expression and inv
RiboTag: A novel technique to profile cell type specific gene expression and inv
批准号:
8473919
负责人:
George STANLEY MCKNIGHT
金额:
$35.95万
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
项目状态:
已结题
起止时间:
2009-09-30 至 2015-05-31
关键词:
AddressAgingAntibodiesBiochemicalBiochemical GeneticsBiological AssayBrainCell NucleusDNA SequenceDiseaseEnvironmentEpitopesEventGene ExpressionGenesGenetic TechniquesGenetic TranslationGoalsHormonesImmunologic TechniquesImmunoprecipitationIn Situ HybridizationIndividualLasersLearningMeasuresMemoryMessenger RNAMethodsMicroarray AnalysisMolecular ProfilingMouse StrainsMusNervous system structureNeuraxisNeuromodulatorNeuronsNeurosciencesNeurotransmittersOccupationsOrganismPharmaceutical PreparationsPhysiologicalPolyribosomesPopulationProtein IsoformsProteinsRNARegulationReporterResolutionReverse Transcriptase Polymerase Chain ReactionRibosomal ProteinsRibosomesSpecificitySpeedSubstance AddictionSubstance abuse problemSynapsesSynaptic plasticityTechniquesTechnologyTestingTimeTransgenic OrganismsTranslatingTranslational Regulationcell typedrug of abuseimprovedin vivonervous system disordernew technologynext generationnovelpost-traumatic stressprotein expressionrecombinaseresponsetooltranscriptome sequencing
中文摘要
项目总结
英文摘要
Project Summary
The brain is composed of hundreds of different neuronal subtypes that each have
unique jobs to do. One of the most significant long-term goals of neuroscience is
to understand how these neuronal subtypes do their job by producing specific
proteins, contacting other neurons, and changing in response to physiological
and pathological contexts. Unfortunately, techniques to capture the total
translated mRNA from a defined subtype of neurons as the organism responds to
the environment, hormones, drugs, and other regulators are lacking. This
proposal addresses that need with a novel biochemical and genetic technique to
express epitope-tagged ribosomal proteins in response to a cell type specific Cre
recombinase in mouse brain. The polyribosomes containing the transcribed and
translatable mRNAs are isolated by immunological techniques and then the RNA
is analyzed by PCR, microarray, and next-generation DNA sequencing. Our
specific aims are to: (1) Develop techniques to optimize polyribosome
immunoprecipitation from brain using the RiboTag mouse that has already been
created with an HA-tagged Rpl22 ribosomal protein under Cre recombinase
regulation. (2) Create a new RiboTag mouse line with a Flag-tagged Rpl23a that
will respond to Cre recombinase activation. (3) Activate Rpl22-HA and Rpl23a-
Flag with specific Cre recombinase transgenics to test the ability of the RiboTag
isolation technique to detect neuron specific mRNAs and the changes that occur
under in vivo physiological regulation. (4) Examine whether the RiboTag
techniques differentiate between translated mRNAs and those that are
translationally repressed. These novel mouse strains and biochemical methods
should allow neuroscientists to measure changes in gene expression and
translational regulation with greater resolution and higher throughput than
previously available and speed our understanding of the underlying changes in
RNA and protein that determine synaptic plasticity.
Relevance
In order to understand how the brain adapts to a changing environment under
physiological circumstances or how specific neuronal populations degenerate or
malfunction in disease, we must have robust and widely available techniques to
measure gene and protein expression in specific neuronal subtypes. The
ribosome tagging (RiboTag) technology we are developing will provide such a
tool to address questions about gene expression and mRNA translation during
adaptive changes in the brain-changes that occur during memory and learning,
substance abuse and addiction, aging, and in response to neurological disorders.
期刊论文(0)
专著(0)
科研奖励(0)
会议论文
Clinical and Basic Studies in Male Reproduction
-
批准号:8065713
-
项目类别:
-
资助金额:$1.0万
-
财政年份:2010
-
负责人:George STANLEY MCKNIGHT
-
依托单位:
Clinical and Basic Studies in Male Reproduction
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批准号:7930074
-
项目类别:
-
资助金额:$24.24万
-
财政年份:2009
-
负责人:George STANLEY MCKNIGHT
-
依托单位:
Clinical and Basic Studies in Male Reproduction
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批准号:7862199
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项目类别:
-
资助金额:$1.0万
-
财政年份:2009
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负责人:George STANLEY MCKNIGHT
-
依托单位:
CAMP AND CALCIUM DEPENDENT KINASES IN SPERMATOGENESIS
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批准号:7553381
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项目类别:
-
资助金额:$40.55万
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财政年份:2007
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负责人:George STANLEY MCKNIGHT
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依托单位:
Protein Kinase A and Intestinal Pseudo-obstruction
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批准号:6704828
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项目类别:
-
资助金额:$15.16万
-
财政年份:2004
-
负责人:George STANLEY MCKNIGHT
-
依托单位:
Protein Kinase A and Intestinal Pseudo-obstruction
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批准号:6896065
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项目类别:
-
资助金额:$15.16万
-
财政年份:2004
-
负责人:George STANLEY MCKNIGHT
-
依托单位:
CAMP AND CALCIUM DEPENDENT KINASES IN SPERMATOGENESIS
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批准号:6588486
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项目类别:
-
资助金额:$15.33万
-
财政年份:2002
-
负责人:George STANLEY MCKNIGHT
-
依托单位:
CAMP AND CALCIUM DEPENDENT KINASES IN SPERMATOGENESIS
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批准号:6655306
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项目类别:
-
资助金额:$15.33万
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财政年份:2002
-
负责人:George STANLEY MCKNIGHT
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依托单位:
ROLE OF CYCLIC AMP DEPENDENT PROTEIN KINASE IN CARDIAC FUNCTION
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批准号:6315351
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项目类别:
-
资助金额:$19.12万
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财政年份:2000
-
负责人:George STANLEY MCKNIGHT
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依托单位:
ROLE OF CAMP AND CALCIUM DEPENDENT PROTEIN KINASES IN SPERMATOGENESIS
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批准号:6311614
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项目类别:
-
资助金额:$16.65万
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财政年份:2000
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负责人:George STANLEY MCKNIGHT
-
依托单位:
ABI PRISM 377 DNA SEQUENCER
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批准号:2803479
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项目类别:
-
资助金额:$13.4万
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财政年份:2000
-
负责人:George STANLEY MCKNIGHT
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依托单位:
ROLE OF CAMP AND CALCIUM DEPENDENT PROTEIN KINASES IN SPERMATOGENESIS
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批准号:6108320
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项目类别:
-
资助金额:$16.65万
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财政年份:1999
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负责人:George STANLEY MCKNIGHT
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依托单位:
ROLE OF CYCLIC AMP DEPENDENT PROTEIN KINASE IN CARDIAC FUNCTION
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批准号:6110047
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项目类别:
-
资助金额:$19.12万
-
财政年份:1999
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负责人:George STANLEY MCKNIGHT
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依托单位:
ROLE OF CAMP AND CALCIUM DEPENDENT PROTEIN KINASES IN SPERMATOGENESIS
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批准号:6296773
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项目类别:
-
资助金额:$16.65万
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财政年份:1999
-
负责人:George STANLEY MCKNIGHT
-
依托单位:
ROLE OF CAMP AND CALCIUM DEPENDENT PROTEIN KINASES IN SPERMATOGENESIS
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批准号:6272020
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项目类别:
-
资助金额:$16.72万
-
财政年份:1998
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负责人:George STANLEY MCKNIGHT
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依托单位:
ROLE OF CYCLIC AMP DEPENDENT PROTEIN KINASE IN CARDIAC FUNCTION
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批准号:6272883
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项目类别:
-
资助金额:$18.36万
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财政年份:1998
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负责人:George STANLEY MCKNIGHT
-
依托单位:
ROLE OF CAMP AND CALCIUM DEPENDENT PROTEIN KINASES IN SPERMATOGENESIS
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批准号:6240875
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项目类别:
-
资助金额:$14.24万
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财政年份:1997
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负责人:George STANLEY MCKNIGHT
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依托单位:
ROLE OF CYCLIC AMP DEPENDENT PROTEIN KINASE IN CARDIAC FUNCTION
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批准号:6242096
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项目类别:
-
资助金额:$17.92万
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财政年份:1997
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负责人:George STANLEY MCKNIGHT
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依托单位:
Regulation of cAMP - Dependent Protein Kinase Genes
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批准号:8272643
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项目类别:
-
资助金额:$39.93万
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财政年份:1997
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负责人:George STANLEY MCKNIGHT
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依托单位:
Regulation of cAMP-Dependent Protein Kinase Genes
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批准号:9066529
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项目类别:
-
资助金额:$46.12万
-
财政年份:1997
-
负责人:George STANLEY MCKNIGHT
-
依托单位:
海外基金