Automated DNA Sequencer and dHPLC Heteroduplex Analysis for Molecular Evolution and Development

摘要9970256基金将用于购买一台Applied Biosystems Prism 377自动DNA测序仪和一台Transgenomics WAVE高效液相色谱仪（dHPLC），供宾夕法尼亚州立大学分子进化遗传学研究所（IMEG）的发育和分子进化遗传学家使用。这些仪器将用于建立一个灵活、高效和具有成本效益的测序和基因分型设施。该策略将采用分层测序方法，使用dHPLC鉴定片段差异并对不同片段进行自动测序。分层方法是一种快速和熟练的方法，用于评估DNA多态性的人口规模的samples.The四个主要研究人员将使用这些工具在一套重叠的项目，整合发育和进化遗传学在广泛的生物体。这些项目包括：与适应性进化相关的基因的分子变异（安德鲁·克拉克）。序列分析将用于探索果蝇中编码代谢酶的基因的调节变化。此外，将进行功能变异和DNA序列多态性的联合分析，以表征和量化果蝇抗菌防御的分子进化和早期发育中的母合子基因表达;第三次染色体倒位中核苷酸多样性的组织（Stephen Schaeffer）。比较DNA序列分析将被用来确定选择，漂移和连锁的相互作用，在确定观察到的遗传变异模式在经典的第三染色体倒位的D。早期发育的遗传调节和与体型中种间差异的关系（比莉·斯瓦拉）。该项目通过识别与海鞘密切相关的Molgula属海鞘物种之间的形态差异的基因来研究身体计划的进化。将在半索动物和尾索动物中确定分子遗传学，并检查可能负责形态进化的候选发育基因;毒力实验进化的遗传基础（托马斯惠特姆）。分子方法使毒力的进化易于在实验室中直接研究。在微生物宿主-寄生虫系统的实验进化制度之后，该项目将比较军团菌的祖先和衍生菌株，并确定用于利用原生动物宿主的已知毒力相关位点的特定核苷酸取代。除了这些主要项目外，IMEG的另外八名活跃成员概述了dHPLC和DNA序列和片段分析的创造性使用。

ABSTRACT 9970256Funds will be used to purchase an Applied Biosystems Prism 377 automated DNA sequencer and a Transgenomics WAVE high performance liquid chromatograph (dHPLC) to be shared by developmental and molecular evolutionary geneticists in the Institute of Molecular Evolutionary Genetics (IMEG) at Penn State University. These instruments will be used to establish a flexible, efficient, and cost-effective sequencing and genotyping facility. The strategy will employ a stratified sequencing approach, using dHPLC to identify fragment differences and automated sequencing of distinct fragments. The stratified approach is a rapid and proficient method for assessing DNA polymorphism in population-sized samples.The four principal investigators will use these instruments in a set of overlapping projects that integrate developmental and evolutionary genetics in a wide range of organisms. The projects include: Molecular variation in genes associated with adaptive evolution (Andrew Clark). Sequence analysis will be used to explore variation in the regulation of genes encoding metabolic enzymes in Drosophila. In addition, combined analysis of functional variation and DNA sequence polymorphism will be performed to characterize and quantify the molecular evolution of antibacterial defense in Drosophila and maternal-zygotic gene expression in early development; Organization of nucleotide diversity in the third chromosomal inversions (Stephen Schaeffer). Comparative DNA sequence analysis will be used to determine the interplay of selection, drift, and linkage in determining observed patterns of genetic variation in the classical third chromosome inversions of D. pseudoobscura; Genetic regulation of early development and associations with interspecific differences in body plans (Billie Swalla). This project is investigating the evolution of body plans by identifying the genes responsible for morphological differences between closely related species of ascidians in the genus Molgula. Molecular phylogenies will be determined and candidate developmental genes that may be responsible for morphological evolution will be examined in hemichordates and urochordates and; Genetic basis of experimental evolution of virulence (Thomas Whittam). Molecular methods render the evolution of virulence amenable to direct study in the laboratory. Following a regime of experimental evolution in a microbial host-parasite system, this project will compare ancestral and derived strains of Legionella and identify specific nucleotide substitutions at known virulence-associated loci used to exploit protozoan hosts. In addition to these major projects, creative use of dHPLC and DNA sequence and fragment analysis is outlined by an additional eight minor users who are active members of IMEG.