BRAIN MORPHOGENESIS DEFECTS IN CEREBUM MUTANT ZEBRAFISH

原肠胚和神经胚期是特别脆弱的 一个是脊椎动物发育，以及遗传和环境扰动 这些过程中的一部分会导致各种先天性畸形 人类神经系统。我们对正常神经和异常神经的理解 系统开发在很大程度上来自于对动物模型的研究 系统。我的最终目标是在动物模型中使用遗传方法 研究脊椎动物神经发育基本过程的系统， 并将我的知识用于培训未来的医生和医务人员 研究人员。在本提案中，我概述了一项综合方案 研究性培训、教学和独立研究，旨在进一步 我的目标。我在卡内基大学攻读博士后培训 机构，一个密集的基础研究环境，在其中我将 学习一套新的动物模型系统和技术。我会有所收获的 有组织的教学经验，通过讲授发育生物学和 对医学和研究生进行小组授课的遗传学课程 在约翰霍普金斯大学医学院，还将参加 利用卡内基研究所提供的机会给予 研讨会和杂志俱乐部。 在我提议的独立研究项目中，我将研究以下方面 硬骨鱼Danio rerio，斑马鱼的早期神经发育。 容易获得的胚胎和精致的基因技术使 斑马鱼是研究潜在遗传规律的一个有吸引力的系统 脊椎动物神经系统的早期发育。一种新的斑马鱼 突变体，大脑(CRB)已被分离出来，基于其显著降低 24小时有脑和眼，尾芽增大。我猜想 CRB的主要缺陷是细胞的会聚运动 原肠形成；因此，细胞被指定为前神经细胞，可能 中胚层结构见于后部。我提议 用胚胎学、遗传学和分子生物学方法鉴定CRB突变 接近了。我将获得CRB的新等位基因，并将使用组合 通过作图和消减杂交分离CRB基因。这就做 对特定的标记进行原位杂交，以评估 中国人神经系统和背侧中胚层的形态和发育 CRB突变体。对细胞行为的延时分析将补充IN 通过产生关于细胞运动的信息进行原位杂交， 繁衍和血统。通过CRB和CRB之间的细胞移植 野生型胚胎我将确定CRB的细胞自主性 行动。最后，我将考察CRB和CRB之间的上位性交互作用 在原肠形成过程中影响细胞运动的其他突变 双重变种人。这一分析应该提供有关 神经板形态发生的基础细胞的行为。

The period of gastrulation and neurulation is a particularly vulnerable one in vertebrate development, and genetic and environmental perturbations of these processes lead to a variety of congenital malformations in the human nervous system. Our understanding of normal and abnormal nervous system development has largely come from the study of animal model systems. My ultimate goal is to use a genetic approach in animal model systems to study fundamental processes in vertebrate neural development, and use my knowledge in the training of future physicians and medical researchers. In the present proposal, I outline an integrated program of research training, teaching, and independent research, designed to further my goals. I am pursuing postdoctoral training at the Carnegie Institution, an intensive basic research environment, in which I will learn a new animal model system and set of techniques. I will gain structured teaching experience, by lecturing in developmental biology and genetics courses that are team taught to the medical and graduate students at the Johns Hopkins University School of Medicine, and will also take advantage of opportunities offered at the Carnegie Institution to give seminars and journal clubs. In my proposed independent research project, I will examine aspects of early neural development in the teleost fish Danio rerio, the zebrafish. Easily accessible embryos and refined genetic techniques make the zebrafish an attractive system to study the genetic regulations underlying early development of the vertebrate nervous system. A new zebrafish mutant, cerebum (crb) has been isolated based on its dramatically reduced brain and eyes at 24 hours, and enlarged tail bud. I hypothesize that the primary defect in crb is in the convergent movement of cells during gastrulation; hence, cells destined for anterior neural and possibly mesodermal structures are found in the posterior region. I propose to characterize the crb mutation using embryologic, genetic, and molecular approaches. I will obtain new alleles of crb, and will use a combination of mapping and subtractive hybridization to isolate the crb gene. I will perform in situ hybridization for specific markers, to evaluate the patterning and development of the nervous system and dorsal mesoderm in crb mutants. Time-lapse analysis of cell behavior will complement the in situ hybridizations by yielding information on cell movements, proliferation, and lineage. Through cell transplants between crb and wild-type embryos I will determine the cell-autonomous nature of crb action. Finally I will examine epistatic interactions between crb and other mutations that effect cell movements in gastrulation by generating double mutants. This analysis should provide new information about the behavior of cells underlying the morphogenesis of the neural plate.