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A Universal Nucleic Acid Recognition Platform for Detection of Pathogenic Bacteria

用于检测病原菌的通用核酸识别平台

基本信息

批准号：
1706802
负责人：
Karin Chumbimuni-Torres
金额：
$ 19.99万
依托单位：
The University of Central Florida Board of Trustees
依托单位国家：
美国
项目类别：
Standard Grant
财政年份：
2017
资助国家：
美国
起止时间：
2017-06-15 至 2021-05-31
项目状态：
已结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=1706802&HistoricalAwards=false
关键词：
Universal Nucleic Acid Recognition Platform

项目摘要

The goal of this research is to develop a low-cost and highly sensitive and selective point-of-care (POC) device that will detect disease-causing bacteria at room temperature. The success of the project requires differentiation between harmless and disease-causing bacteria, and is possible by detection of tiny differences in bacterial RNA (genetic material). The proposed research presents a sensor device with a simple design that consists of a universal nucleic acid probe and two intermediary strands of nucleic acids organized in such a way that changes in their natural association can be used to measure the presence the disease-causing bacteria. Broader impacts for education and the community include the development of the outreach program called GRoW uP in the Central Florida area.Current approaches for detecting pathogens, such as nucleic acid amplification tests (NAATs), offer high sensitivity, but often produce false-positive results, and require expensive instruments and trained personnel. Furthermore, these approaches cannot differentiate between the point mutations that distinguish the RNA of the pathogenic from the non-pathogenic bacteria. This is especially difficult at ambient temperatures and for RNA that forms secondary structure. The goal of this proposal is to develop a specific and sensitive universal point-of-care (POC) platform for the rapid and accurate detection of pathogenic bacteria. The approach uses a binary probe with high sequence recognition specificity that is necessary to differentiate point mutations and is amenable to analysis of folded RNA. The new methodology will exceed the performance of current state-of-the-art approaches for RNA sensing in the following aspects: (i) it will exhibit zero false-negative responses, thus improving sensitivity and reducing limits of detection (LOD); (ii) it will enable accurate recognition of point mutations at ambient temperatures even in structured (folded) RNA and DNA; (iii) it will enable detection of multiple analytes using a single universal probe in a re-usable format and thus reduce the assay cost. The proposed work builds upon the PI?s recent improvements on the self-assembling DNA 4J fluorescent sensor by converting the system to an electrochemical biosensor (4J E-biosensor). Advantages of the 4J E-biosensor include the capability of performing multiple tests with zero signal background and the potential for on-site testing while offering fast, simple, low-power, portable, and inexpensive detection. The novel sensor platform will advance the ability to detect ribosomal RNA and thus, have unprecedented impact in environmental, health diagnosis, national defense and food safety applications. The interdisciplinary nature of the proposed work will expose students to different areas of chemistry, including electrochemistry, biochemistry, computational simulations, sensing applications, and nanomaterial sciences. The PI proposes to use this research as a motivation to continue and expand her self-developed GRoW uP program. The program will provide research opportunities to high school students and teachers through partnerships established with Orange Public School District. The PI will also implement an outreach program in conjunction with the Orlando Science Center and the Seminole County Library to impact hundreds of young people across multi-ethnic Orlando and Central Florida. Dissemination of the results from this investigation will be through publications in high-impact peer-reviewed journals and national and international conferences.

这项研究的目标是开发一种低成本、高灵敏度和选择性的护理点(POC)设备，该设备将在室温下检测致病细菌。该项目的成功需要区分无害细菌和致病细菌，通过检测细菌RNA(遗传物质)的微小差异是可能的。这项拟议的研究提出了一种设计简单的传感器设备，它由一个通用的核酸探针和两条中间核酸组成，其组织方式是，它们自然结合的变化可以用来测量致病细菌的存在。对教育和社区的更广泛影响包括发展一项名为在佛罗里达中部地区成长的推广计划。目前检测病原体的方法，如核酸扩增测试(NAAT)，具有很高的灵敏度，但经常产生假阳性结果，并且需要昂贵的仪器和训练有素的人员。此外，这些方法不能区分致病细菌和非致病细菌的RNA点突变。这在环境温度和形成二级结构的RNA中尤其困难。这项提议的目标是开发一个特定和敏感的通用护理点(POC)平台，用于快速和准确地检测病原菌。该方法使用具有高序列识别特异性的二元探针，这是区分点突变所必需的，并且适合于折叠RNA的分析。新方法将在以下方面超过目前最先进的RNA检测方法的性能：(I)它将表现出零假阴性反应，从而提高灵敏度并降低检测限(LOD)；(Ii)它将能够在常温下准确识别点突变，即使是在结构(折叠)的RNA和DNA中；(Iii)它将能够以可重复使用的格式使用单个通用探针检测多个分析物，从而降低分析成本。这项工作建立在PiooS最近对自组装DNA4J荧光传感器的改进上，将该系统转换为电化学生物传感器(4JE-生物传感器)。4J E-生物传感器的优势包括能够在零信号背景下执行多个测试，并有可能进行现场测试，同时提供快速、简单、低功耗、便携和廉价的检测。这种新型的传感器平台将提高检测核糖体RNA的能力，从而在环境、健康诊断、国防和食品安全应用中产生前所未有的影响。拟议工作的跨学科性质将使学生接触不同的化学领域，包括电化学、生物化学、计算模拟、传感应用和纳米材料科学。PI建议将这项研究作为继续和扩大她自己开发的成长计划的动力。该计划将通过与奥兰治公立学区建立合作伙伴关系，为高中生和教师提供研究机会。PI还将与奥兰多科学中心和塞米诺尔县图书馆一起实施一项外联计划，以影响奥兰多和佛罗里达州中部多民族地区的数百名年轻人。将通过在影响较大的同行评议期刊以及国内和国际会议上发表出版物来传播这项调查的结果。

项目成果

期刊论文数量（8）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

A universal and label-free impedimetric biosensing platform for discrimination of single nucleotide substitutions in long nucleic acid strands

DOI：
10.1016/j.bios.2018.02.059
发表时间：
2018-06-30
期刊：
BIOSENSORS & BIOELECTRONICS
影响因子：
12.6
作者：
Mills, Dawn M.;Martin, Christopher P.;Chumbimuni-Torres, Karin Y.
通讯作者：
Chumbimuni-Torres, Karin Y.

MVF Sensor Enables Analysis of Nucleic Acids with Stable Secondary Structures

DOI：
10.1002/elan.201900690
发表时间：
2020-01
期刊：
Electroanalysis
影响因子：
3
作者：
M. Foguel;Angelica M. Balcarcel;M. Reed;Percy Calvo-Marzal;Y. Gerasimova;D. Kolpashchikov;K. Chumbimuni-Torres
通讯作者：
M. Foguel;Angelica M. Balcarcel;M. Reed;Percy Calvo-Marzal;Y. Gerasimova;D. Kolpashchikov;K. Chumbimuni-Torres

An electrochemical biosensor based on Hairpin-DNA modified gold electrode for detection of DNA damage by a hybrid cancer drug intercalation

DOI：
10.1016/j.bios.2019.02.071
发表时间：
2019-05-15
期刊：
BIOSENSORS & BIOELECTRONICS
影响因子：
12.6
作者：
Untiveros, Katherine Lozano;da Silva, Emanuella Gomes;Chumbimuni-Torres, Karin
通讯作者：
Chumbimuni-Torres, Karin

Selective Determination of Isothermally Amplified Zika Virus RNA Using a Universal DNA-Hairpin Probe in Less than 1 Hour