CRISPR interference-enabled phenotyping of essential genes in C. difficile to aid in discovery of antibiotic targets
对艰难梭菌中的必需基因进行 CRISPR 干扰表型分析,以帮助发现抗生素靶标
基本信息
- 批准号:10518406
- 负责人:
- 金额:$ 17.38万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2021
- 资助国家:美国
- 起止时间:2021-11-02 至 2024-10-31
- 项目状态:已结题
- 来源:
- 关键词:AffectAntibiotic ResistanceAntibioticsBacillus subtilisBacteriaBacterial GenesBioinformaticsBiological AssayCRISPR interferenceCategoriesCell WallCell divisionCell physiologyCellsCessation of lifeCloningClostridiumClostridium difficileCommunitiesCytolysisDNA SequenceDNA biosynthesisDNA-Directed RNA PolymeraseDefectDevelopmentDrug InteractionsDrug TargetingEmerging TechnologiesEscherichia coliEssential GenesFailureFilamentFluorescence MicroscopyFutureGene ClusterGene ExpressionGenesGenetic TranscriptionGuide RNAHealthHospitalsHumanIndividualInfectionKnowledgeLeadLibrariesLinkLipidsMetabolismMethodsMorphologyMutagenesisNursing HomesOperonOrganismOrthologous GenePathway interactionsPersonsPharmaceutical PreparationsPhasePhenotypePhysiologicalPlasmidsProcessProtein InhibitionProteinsProtocols documentationPublic HealthQuantitative Reverse Transcriptase PCRRNARepressionResearchResearch PersonnelResourcesRodRoleShapesSystemTechniquesTranslationsUnited StatesWorkXylosedeep sequencingexperimental studyfollow-upgene functiongene repressionglycosyltransferasegut microbiotaimprovedinnovationinsightinterestknock-downmicrobiotanew therapeutic targetparalogous genepreventpromoterprotein-histidine kinaserepositoryscale upscreeningsegregationsynergismsynthetic constructtherapy outcometooltransposon sequencing
项目摘要
Project Summary
Clostridioides (Clostridium) difficile infections strike close to 500,000 people a year in the United States, leading
to nearly 30,000 deaths. The CDC has declared this organism an “urgent” threat to public health, the highest
threat category. New treatments are sorely needed. Most antibiotics inhibit proteins that are essential for viability,
so a better understanding of these proteins and the genes that encode them can advance efforts to develop new
antibiotics. Although essential genes are difficult to study, new techniques for manipulating bacterial gene
expression are overcoming this barrier. One emerging technology for studying essential genes in bacteria is
called CRISPR interference (CRISPRi). CRISPRi uses a short RNA to guide a protein called dCas9 to a gene,
thereby creating a roadblock that prevents RNA polymerase from transcribing that gene. CRISPRi is well-suited
for screening large sets of genes because repressing a new target is simply a matter of cloning a synthetic DNA
sequence encoding the right guide RNA into the plasmid, a process that is easily scaled-up. Another key feature
of our CRISPRi system is that dCas9 is expressed from a xylose-inducible promoter, which allows us to control
the timing and extent of gene repression. The specific objective of this proposal is to deploy CRISPRi to
characterize 187 putatively essential genes in C. difficile. In Aim 1 we will construct a CRISPRi library and use it
to vet the essentiality of the genes on our target list. In Aim 2 we will determine whether gene knockdown results
in morphological defects or increases sensitivity to a panel of antibiotics chosen to target a variety of cellular
processes. The information from these assays will enable us to assign many of these genes to functional
pathways. It will also provide foundational knowledge and tools for developing new antibiotics to treat C. difficile
infections.
项目摘要
在美国,艰难梭菌感染每年造成近50万人死亡,居首位
造成近3万人死亡。美国疾病控制与预防中心宣布这种微生物是对公共健康的“紧急”威胁,这是最高的
威胁类别。迫切需要新的治疗方法。大多数抗生素会抑制对生存至关重要的蛋白质,
因此,更好地了解这些蛋白质和编码它们的基因可以推动开发新的
抗生素。尽管基本基因很难研究,但操纵细菌基因的新技术
表达正在克服这一障碍。研究细菌基本基因的一项新兴技术是
称为CRISPR干扰(CRISPRi)。CRISPRi使用一种短RNA来引导一种名为dCas9的蛋白质进入基因,
从而造成了一个障碍,阻止了RNA聚合酶转录该基因。CRISPRi非常适合
用于筛选大组基因,因为抑制一个新的靶点只需克隆合成的DNA
将编码正确的引导RNA的序列插入到质粒中,这一过程很容易放大。另一个关键功能
我们CRISPRi系统的一个特点是,dCas9是由木糖诱导的启动子表达的,这使得我们能够控制
基因抑制的时间和程度。这项提议的具体目标是将CRISPRi部署到
艰难梭菌187个假定必需基因的特征。在目标1中,我们将构建一个CRISPRi库并使用它
来检查我们目标名单上的基因的重要性。在目标2中,我们将确定基因敲除结果
在形态上的缺陷或增加对一组被选择来针对各种细胞的抗生素的敏感性
流程。来自这些分析的信息将使我们能够将这些基因中的许多分配到功能上
小路。它还将为开发治疗艰难梭菌的新抗生素提供基础知识和工具。
感染。
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(0)
数据更新时间:{{ journalArticles.updateTime }}
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
数据更新时间:{{ journalArticles.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ monograph.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ sciAawards.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ conferencePapers.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ patent.updateTime }}
DAVID S WEISS其他文献
DAVID S WEISS的其他文献
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
{{ truncateString('DAVID S WEISS', 18)}}的其他基金
Heteroresistance Interdisciplinary Research Unit (Project 2)
异阻性跨学科研究单元(项目2)
- 批准号:
10366038 - 财政年份:2021
- 资助金额:
$ 17.38万 - 项目类别:
CRISPR interference-enabled phenotyping of essential genes in C. difficile to aid in discovery of antibiotic targets
对艰难梭菌中的必需基因进行 CRISPR 干扰表型分析,以帮助发现抗生素靶标
- 批准号:
10369416 - 财政年份:2021
- 资助金额:
$ 17.38万 - 项目类别:
Heteroresistance Interdisciplinary Research Unit (Project 2)
异阻性跨学科研究单元(项目2)
- 批准号:
10583505 - 财政年份:2021
- 资助金额:
$ 17.38万 - 项目类别:
Heteroresistance Interdisciplinary Research Unit (Project 2)
异阻性跨学科研究单元(项目2)
- 批准号:
10170971 - 财政年份:2021
- 资助金额:
$ 17.38万 - 项目类别:
Heteroresistance Interdisciplinary Research Unit (Admin Core)
异阻性跨学科研究单位(行政核心)
- 批准号:
10170967 - 财政年份:2021
- 资助金额:
$ 17.38万 - 项目类别:
Heteroresistance Interdisciplinary Research Unit (Admin Core)
异阻性跨学科研究单位(行政核心)
- 批准号:
10583498 - 财政年份:2021
- 资助金额:
$ 17.38万 - 项目类别:
Heteroresistance Interdisciplinary Research Unit (Admin Core)
异阻性跨学科研究单位(行政核心)
- 批准号:
10366034 - 财政年份:2021
- 资助金额:
$ 17.38万 - 项目类别:
Exploitation of multiple heteroresistance for effective antibiotic combination therapy
利用多重异质耐药性进行有效的抗生素联合治疗
- 批准号:
10646392 - 财政年份:2020
- 资助金额:
$ 17.38万 - 项目类别:
Exploitation of multiple heteroresistance for effective antibiotic combination therapy
利用多重异质耐药性进行有效的抗生素联合治疗
- 批准号:
10206015 - 财政年份:2020
- 资助金额:
$ 17.38万 - 项目类别:
Exploitation of multiple heteroresistance for effective antibiotic combination therapy
利用多重异质耐药性进行有效的抗生素联合治疗
- 批准号:
10053046 - 财政年份:2020
- 资助金额:
$ 17.38万 - 项目类别:
相似海外基金
The effects of antibiotics to the transfer frequency of the antibiotic resistance genes and the evolution of high-level resistance.
抗生素对抗生素抗性基因转移频率和高水平抗性进化的影响。
- 批准号:
22K05790 - 财政年份:2022
- 资助金额:
$ 17.38万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
NEC05839 Chicken or the Egg: Is AMR in the Environment Driven by Dissemination of Antibiotics or Antibiotic Resistance Genes?
NEC05839 先有鸡还是先有蛋:环境中的抗菌素耐药性是由抗生素或抗生素抗性基因的传播驱动的吗?
- 批准号:
NE/N019687/2 - 财政年份:2019
- 资助金额:
$ 17.38万 - 项目类别:
Research Grant
Combating Antibiotic Resistance to Aminoglycoside Antibiotics through Chemical Synthesis
通过化学合成对抗氨基糖苷类抗生素的耐药性
- 批准号:
392481159 - 财政年份:2017
- 资助金额:
$ 17.38万 - 项目类别:
Research Fellowships
NEC05839 Chicken or the Egg: Is AMR in the Environment Driven by Dissemination of Antibiotics or Antibiotic Resistance Genes?
NEC05839 先有鸡还是先有蛋:环境中的抗菌素耐药性是由抗生素或抗生素抗性基因的传播驱动的吗?
- 批准号:
NE/N019687/1 - 财政年份:2016
- 资助金额:
$ 17.38万 - 项目类别:
Research Grant
Chicken or the Egg: Is AMR in the Environment Driven by Dissemination of Antibiotics or Antibiotic Resistance Genes?
先有鸡还是先有蛋:环境中的抗菌素耐药性是由抗生素或抗生素抗性基因的传播驱动的吗?
- 批准号:
NE/N019857/1 - 财政年份:2016
- 资助金额:
$ 17.38万 - 项目类别:
Research Grant
The SuDDICU study- A study of the impact of preventative antibiotics (SDD) on patient outcome and antibiotic resistance in the critically ill in intensive care
SuDDICU 研究 - 一项关于预防性抗生素 (SDD) 对重症监护病危患者的患者预后和抗生素耐药性影响的研究
- 批准号:
366555 - 财政年份:2016
- 资助金额:
$ 17.38万 - 项目类别:
Operating Grants
Chicken or the Egg: Is AMR in the Environment Driven by Dissemination of Antibiotics or Antibiotic Resistance Genes?
先有鸡还是先有蛋:环境中的抗菌素耐药性是由抗生素或抗生素抗性基因的传播驱动的吗?
- 批准号:
NE/N019717/1 - 财政年份:2016
- 资助金额:
$ 17.38万 - 项目类别:
Research Grant
The SuDDICU study- A study of the impact of preventative antibiotics (SDD) on patient outcome and antibiotic resistance in the critically ill in intensive care
SuDDICU 研究 - 一项关于预防性抗生素 (SDD) 对重症监护病危患者的患者预后和抗生素耐药性影响的研究
- 批准号:
361307 - 财政年份:2016
- 资助金额:
$ 17.38万 - 项目类别:
Operating Grants
Contamination status of antibiotics and antibiotic resistance genes (ARGs) in tropical Asian aquatic environments with artificial and natural disturbance
人工和自然干扰下亚洲热带水生环境中抗生素和抗生素抗性基因(ARG)的污染状况
- 批准号:
25257402 - 财政年份:2013
- 资助金额:
$ 17.38万 - 项目类别:
Grant-in-Aid for Scientific Research (A)
RAPID: COLLABORATIVE RESEARCH: Fate and Transport of Antibiotics and Antibiotic Resistance Genes During Historic Colorado Flood
快速:合作研究:历史性科罗拉多洪水期间抗生素和抗生素抗性基因的命运和运输
- 批准号:
1402635 - 财政年份:2013
- 资助金额:
$ 17.38万 - 项目类别:
Standard Grant