Acquisition of an Automated DNA Sequencer for The New York Botanical Garden's Cullman Program for Molecular Systematics Studies

为纽约植物园 Cullman 分子系统学研究项目购置自动 DNA 测序仪

• 批准号: 9601515
• 负责人: Victor Albert
• 金额: $ 0.93万
• 依托单位: New York Botanical Garden
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-09-01 至 1997-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9601515&HistoricalAwards=false
• 关键词: Acquisition; Automated; DNA; Sequencer; New

This proposal seeks cost-sharing support for the purchase of an ABI PRISM 377 Automated DNA Sequencer and necessary peripherals. The Lewis B. and Dorothy Cullman Program for Molecular Systematics Studies, which is a joint initiative of The New York Botanical Garden and the American Museum of Natural History, was formed through a charitable donation from the Lewis B. and Dorothy Cullman Foundation. The New York Botanical Garden's Cullman Program occupies laboratories at both locations, which includes entitlement to user-fee based operation of the Museum's ABI 377 sequencer. The Cullman Program resides primarily in newly renovated laboratory space in the Garden's Harding Laboratory building. A matching challenge grant of $116,000 from the Cullman Foundation has been used toward the total costs of equipping the new laboratories for molecular systematics research. This proposal requests the balance of necessary funds from the NSF. This proposal seeks matching funds from NSF to complete the planned equipping of the Garden laboratories with automated DNA sequencing and DNA fingerprinting capacities. DNA sequencing is not currently possible at the Garden, although all other relevant molecular techniques (e.g., DNA and RNA extraction and purification, Southern and northern blot analyses, PCR amplification of DNA, in situ hybridization and in situ PCR against DNA or cDNA, visualization of fluorochrome labeled in situ probes, and molecular cloning) are supported with the newly purchased equipment. The sequencing facility at the American Museum is crucial for present research initiatives, but it is no long term substitute for in-house DNA sequence and DNA fingerprint determination at the Garden. Aspects such as cost-effectiveness, throughput, flexibility, and enhanced research and educational development argue strongly for the Garden's completion of its own automated DNA analysis facility, as has been planned. The general goal of the Cullman Program is to enr ich and invigorate the Botanical Science Division of The New York Botanical Garden with the modern tools of molecular systematics research. Molecular systematics not only entails DNA sequence production for use in phylogenetic analysis; it also encompasses bold new attempts to link systematics reasoning with the burgeoning field of plant molecular development. The Cullman Program currently supports standard molecular systematics projects as well as systematically oriented projects focusing on flower and root development. Molecular systematic work in Lecythidaceae, Arecaceae, Anacardiaceae, Neckeraceae, Gentianaceae, and Cactaceae is proceeding in collaboration with Cullman Program Adjunct Curators, other staff, and graduate students. These projects are primarily oriented toward classificatory, character evolutionary, and biogeographic issues. Other projects focusing on population level issues will profit from the ABI 377's capacity to perform rapid genotype analysis (e.g., using the amplified fragment length polymorphism AFLP approach). Likewise, the evolutionary-oriented developmental research of the Cullman Program requires active DNA sequencing capacity. A Garden sequencing facility will act much like a local workstation for plant molecular systematics activity, especially given the Garden's academic collaborations with Columbia University (The Center for Environmental Conservation and Research--CERC), Cornell University, Lehman College--CUNY, New York University, and Yale University. Immediate commitments for use of an automated sequencer are substantial (16 investigators), and installation of this instrumentation at The New York Botanical Garden should permit a considerable molecular systematics research capacity in the years to come. The ABI PRISM 377 Automated DNA Sequencer (manufactured by Perkin Elmer's Applied Biosystems Division) is a sophisticated instrument representing the forefront of second-generation sequencer technology. The instrument' s use of simultaneous, scanning-laser detection of four different fluorescent dyes permits samples to be resolved in single gel lanes (as opposed to the four gel lanes occupied by other vendors' equipment, which have a single-dye approach). Throughput time has been increased by a factor four from earlier models through refinements to the scanning procedure. Combined with appropriate software (some software and hardware are included within the base purchase price), gel-plate and loading comb combinations, apparatus for PCR-based DNA labeling, and a dedicated water purification system, the ABI 377 will complete the equipping of The New York Botanical Garden's molecular laboratories.

这项提案寻求为购买ABI PRISM 377自动DNA测序仪和必要的外围设备提供成本分摊支持。刘易斯·B·卡尔曼和多萝西·卡尔曼分子系统学研究计划是由纽约植物园和美国自然历史博物馆联合发起的，通过刘易斯·B和多萝西·卡尔曼基金会的慈善捐赠而成立。纽约植物园的卡尔曼计划占据了这两个地点的实验室，其中包括有权使用博物馆的ABI 377测序仪进行收费操作。卡尔曼计划主要位于花园哈丁实验室大楼内新装修的实验室空间内。卡尔曼基金会提供的11.6万美元的等额挑战赠款已用于装备新实验室进行分子系统学研究的总成本。这项提案要求国家科学基金会平衡必要的资金。这项提案寻求国家科学基金会提供配套资金，以完成为花园实验室配备自动DNA测序和DNA指纹能力的计划。花园目前还不能进行DNA测序，尽管所有其他相关的分子技术(例如DNA和RNA的提取和纯化、Southern和Northern印迹分析、DNA的聚合酶链式反应扩增、针对DNA或cDNA的原位杂交和原位聚合酶链式反应、荧光标记的原位探针可视化以及分子克隆)都得到了新购买的设备的支持。美国博物馆的测序设施对目前的研究活动至关重要，但它不能长期取代花园的内部DNA序列和DNA指纹测定。在成本效益、吞吐量、灵活性以及加强的研究和教育发展等方面，都强烈支持花园按照计划完成自己的自动化DNA分析设施。卡尔曼计划的总体目标是用分子系统学研究的现代工具激励和振兴纽约植物园的植物科学部。分子系统学不仅包括用于系统发育分析的DNA序列生产；它还包括将系统学推理与新兴的植物分子发展领域联系起来的大胆的新尝试。卡尔曼计划目前支持标准的分子系统学项目，以及侧重于花和根发育的系统导向项目。Lecythidaceae、Arecaceae、Anacardiaceae、Neckeraceae、Gentianaceae和Cactaceae的分子系统工作正在与卡尔曼计划助理馆长、其他工作人员和研究生合作进行。这些项目主要针对分类、特征进化和生物地理问题。其他专注于种群水平问题的项目将受益于ABI377‘S进行快速基因分析的能力(例如，使用扩增片段长度多态性AFLP方法)。同样，卡尔曼计划以进化为导向的发展研究需要活跃的DNA测序能力。花园的测序设施将非常像植物分子系统学活动的本地工作站，特别是考虑到花园与哥伦比亚大学(环境保护与研究中心-CERC)、康奈尔大学、雷曼学院-纽约大学、纽约大学和耶鲁大学的学术合作。立即承诺使用自动测序仪是很重要的(16名研究人员)，在纽约植物园安装这种仪器应该会在未来几年允许相当大的分子系统学研究能力。ABI PRISM 377自动DNA测序仪(由Perkin Elmer应用生物系统事业部制造)是一种代表第二代测序仪技术前沿的尖端仪器。该仪器的S使用四种不同荧光染料的同时扫描激光检测，允许在单一凝胶通道中解析样品(而不是其他供应商的设备占用四个凝胶通道，后者采用单一染料方法)。通过对扫描程序的改进，吞吐时间比以前的型号增加了四倍。结合适当的软件(一些软件和硬件包含在基本购买价格中)、凝胶板和加载梳子组合、基于PCR的DNA标记设备和专用水净化系统，ABI 377将完成纽约植物园分子实验室的配备。