Methods for Detecting Nucleic Acid Hybridization Using Liquid Crystals

CBET-1160202 Schwartz该项目的总体目标涉及开发使用液晶（LC）检测核酸杂交的方法。此外，为了获得一个基本的分子水平的机制，其中核酸的界面相互作用影响LC取向的理解。本研究的最终长期目标是使基于LC的检测在医学诊断应用中成为可能，例如基于LC对DNA杂交的取向响应的多路复用方法（即DNA微阵列），该方法使用偏振光的透射来检测LC响应。第一种方法涉及在阳离子表面活性剂单层存在下LC/水界面处LC极性倾斜角的动态变化，与该方法相关的具体目标包括开发多路复用平台，定量研究灵敏度和目标通用性，开发可用于研究LC/水界面的荧光显微镜方法，以及这些方法在理解LC响应DNA杂交的分子机制中的应用。最后，该方法将通过实验直接验证，该实验将区分彼此相关的细菌病原体。第二种方法涉及平面-当LC与延伸的dsDNA接触时锚定LC。相关的specificaims包括定量研究的宏观光学响应，扩展的方法，以检测单个杂交分子，开发的方法，使多路检测使用共价固定的探针，和机制的研究，旨在了解不同的LC响应ssDNA和dsDNA。类似的验证实验（即相关细菌之间的歧视）将追求这种方法作为第一个方法。更广泛的影响这里开发的方法具有长期的潜力，可用于新的医疗诊断技术，适用于使用点的应用，包括那些在低资源环境。此外，对LC-生物分子相互作用的基本理解将对研究人员追求基于LC的生物传感方法广泛有用。所开发的光学表征方法将用于LC-液体界面的研究以及其它液体-液体界面。参与该项目的研究人员将获得多学科领域的专业知识，预计将在未来的科学和技术的前沿几十年来，包括液晶器件，生物检测，单分子显微镜和自组装分子系统。此外，通过参与各种教育和推广计划，（例如REU，RET，GAANN，CU-发现学习中心，CU K-12外展材料科学，科罗拉多高中荣誉研究所），研究人员将继续与学生，教师，和其他社区成员。正如提案中所总结的那样，PI和参与该项目的前学生和博士后已经表现出真诚和持续的承诺，创造性地利用他们的研究来追求教育和推广;该项目中基本发现和尖端显微镜方法的整合提供了一个理想的平台，从这个平台向公众传达研究的兴奋。

CBET-1160202SchwartzThe overall objectives of this project involve the development of methods where liquid crystals(LC) are used to detect nucleic acid hybridization. Also, to obtain an improved understanding ofthe fundamental molecular-level mechanisms by which the interfacial interactions of nucleicacids affect LC orientation. The ultimate long-term goal of this research is to enable LC-baseddetection in medical diagnostic applications, e.g. multiplexed methods (i.e. DNA microarrays)based on the orientational response of LCs to DNA hybridization, that use the transmission ofpolarized light to detect the LC response.Intellectual MeritTwo distinct approaches will be developed, and so the specific aims and research strategies ofthis project fall into two groups. The first approach involves a dynamic change of the LC polar tiltangle at the LC/aqueous interface in the presence of a monolayer of cationic surfactant, and thespecific aims associated with this approach include the development of a multiplexed platform,quantitative studies of sensitivity and target generality, the development of fluorescencemicroscopy methods that can be used to study the LC/aqueous interface, and the application ofthese methods to understand the molecular mechanisms of LC response to DNA hybridization.Finally, this method will be directly validated by experiments that will discriminate betweenclosely-related bacterial pathogens.The second approach involves a characteristic chiral rotation of the azimuthal orientation ofplanar-anchored LC when the LC is in contact with extended dsDNA. The associated specificaims include quantitative studies of the macroscopic optical response, extension of the methodto the detection of single hybridized molecules, development of methods that enable multiplexeddetection using covalently immobilized probes, and mechanistic studies aimed at understandingthe distinctive LC response to ssDNA and dsDNA. Similar validation experiments (i.e.discrimination between related bacteria) will be pursued for this method as for the first method.Broader ImpactThe methods developed here have the long-term potential to be used in new medical diagnostictechnologies that are appropriate for point-of-use applications including those in low resourceenvironments. Furthermore, the fundamental understandings obtained about LC-biomoleculeinteractions will be broadly useful for researchers pursuing LC-based biosensing approaches.More broadly, the optical characterization methods developed will be of use for studies of theLC-liquid interface and for other liquid-liquid interfaces.The researchers involved in this project will gain expertise in multidisciplinary areas projected tobe at the forefront of science and technology in the coming decades, including liquid crystaldevices, biodetection, single-molecule microscopy, and self-assembled molecular systems.Also, through their participation in a variety of educational and outreach programs (e.g. REU,RET, GAANN, CU-Discovery Learning Center, Materials Science from CU K-12 outreach,Colorado High School Honors Institute), the researchers will continue to share this project withstudents, teachers, and other community members. As summarized in the proposal the PI andprevious students and post-docs involved in this project have demonstrated a sincere andcontinuing commitment to the creative use of their research in the pursuit of education andoutreach; the integration of basic discovery and cutting-edge microscopy methods in this projectwill provide an ideal platform from which to communicate the excitement of research to thegeneral public.