Aquisition of a DNA Sequencer for Plant Molecular Biology Research

目前植物分子生物学研究的基础是对具有生物学和经济学意义的基因进行常规测序的能力。 这些基因正在从高等植物物种以及作为植物共生体和病原体的许多微生物中克隆为基因组和cDNA片段。 分离拟南芥突变体和测序特定改变表型的基因的能力，为破译植物生长和发育的分子机制提供了一种强有力的新手段。 不幸的是，研究生和博士后同事在植物分子生物学实验室中花费的大部分实际研究时间都被机械、费力的程序占用了，通过这些程序，DNA测序“阶梯”被精确地分离、检测和分析。 我们估计，他们至少有30%的时间被这个过程所占用，这需要时间来进行更关键和更严格的生物实验设计。 这项提案的目的是获得联邦配套资金，以购买自动DNA测序仪。 该机器将由Sussman教授监督，并由威斯康星州基金提供工资的全职技术人员在其实验室内操作。 这些自动化的基于荧光的测序仪广泛用于基因组测序项目，并且已经被证明是节省时间和劳动力的成本有效的手段。 除了节省时间外，自动化技术的其他优点包括没有放射性核素，以及减少与其安全和处置有关的财政负担。 该设备的主要用户包括8个在植物分子生物学领域有联邦资助基础研究计划的教师。 这些学院的实验室在物理上非常接近，但通过5个不同的学术部门（园艺，农学，生物化学和植物病理学在农业和生命科学学院的部门;植物学在文学和科学学院的部门）进行管理。 少数用户包括几名新雇用的“应用”遗传学家，他们正在开展利用重组DNA技术更有效地培育改良作物的研究方案。

Underpinning much of the current research in plant molecular biology is the ability to routinely sequence genes of biological and economic interest. These genes are being cloned as genomic and cDNA fragment from higher plant species, as well as from the many microbes that act as plant symbionts and pathogens. The ability to isolate Arabidopsis mutants and sequence the genes underlying specific altered phenotypes has introduced a powerful new means of deciphering the molecular mechanisms by which plants grow and develop. Unfortunately, much of the actual research time spent by graduate students and postdoctoral associates in plant molecular biology laboratories is occupied with the rote, laborious procedure by which DNA sequencing `ladders' are electrophoretically separated, detected and analyzed. We estimate that at least 30% of their time is occupied with this procedure, which takes time away from the more critical and intellecutually demanding design of biological experiments. The purpose of this proposal is to obtain federal matching funds to purchase an automated DNA sequencer. This machine will be supervised by Prof. Sussman and operated in his laboratory by a full-time technician with salary provided by state of Wisconsin funds. These automated, fluorescent- based sequencers are widely used in genome sequencing projects, and have been shown to be a cost-effective means of saving time and labor. In addition to time savings, other advantages of the automated technique include the absence of radioactive nuclides and a decreased financial burden associated with their safety and disposal. Major users of this equipment include 8 faculty with federally funded basic research program s in the area of plant molecular biology. The laboratories of these faculty are in close physical proximity but are administered through 5 different academic departments (Departments of Horticulture, Agronomy, Biochemistry and Plant Pathology in the College of Agricultural and Life Sciences; Department of Botany in the College of Letters and Science). Minor users include several newly hired `applied' geneticists with active research programs using recombinant DNA techniques for more efficient breeding of improved crop plants.