CSHL Cell and Developmental Biology of Xenopus

描述(由申请人提供)：近年来，细胞和发育生物学领域取得了巨大的进步，这主要是由于对经典问题的新技术方法的应用。这导致了对高级课程的需求，以培训研究人员研究那些在新发现中发挥重要作用的动物模型，如非洲爪哇。目前正在寻求资金，以继续开设冷泉港实验室课程“非洲爪哇的细胞和发育生物学”，该课程作为现代生物学若干领域综合课程的一部分，已教授了11年。本课程提供对非洲爪蛙和热带非洲爪蛙胚胎的生物学和操作的广泛实验室接触。非洲爪哇是一种唯一适合研究早期发育的脊椎动物，因为从受精开始，很容易获得大量的胚胎。发育速度很快，胚胎的大小有利于他们的微操作。这使得可以分析(如果必要的话，在单细胞水平上)在哺乳动物、鸟类或鱼类胚胎中不可行的母体调节和胚胎诱导等重要问题。这门课程既适合那些有分子生物学和发育生物学知识但没有使用非洲爪哇经验的人，也适合那些有一些非洲爪哇经验但希望学习新开发的先进技术的人。具体内容包括：(1)非洲爪哇成虫的护理和处理、去除卵母细胞、诱导排卵、收集卵子、分离睾丸、体外受精和自然受精；(2)胚胎发育和解剖阶段；(3)整体原位杂交和免疫组织化学，包括双探针染色；(4)用谱系示踪剂、反义寡核苷酸、信使核糖核酸和DNA结构显微注射卵和卵母细胞；(5)胚胎的显微操作，包括诱导和移植试验；(6)用于母体分子“敲除”的寡核苷酸反义技术；(7)用于构建转基因胚胎的细胞周期提取物的制备和使用；(8)使用热带非洲爪哇作为遗传模型。 预计在拟议的支助期内，课程的基本安排和大部分课程材料将保持不变。然而，快速发展的主题，如在体内使用siRNA进行基因敲除，用于转基因的新启动子序列，以及转基因和突变株的可用性，将随着该领域的进展而被讨论和整合到课程中。

DESCRIPTION (provided by applicant): Dramatic progress has been made in the fields of cell and developmental biology in recent years, largely due to the application of new technical approaches to classical problems. This has led to the need for advanced courses to train investigators to study those animal models, such as Xenopus, that have been important in making new discoveries. Funds are being sought to continue the Cold Spring Harbor Laboratory course on "Cell and Developmental Biology of Xenopus" which has been taught for eleven years as part of an integrated set of courses in a number of areas of modern biology. This course provides extensive laboratory exposure to the biology and manipulation of embryos from the frogs Xenopus laevis and Xenopus tropicalis. Xenopus is a vertebrate uniquely suited for studies of early development, since large numbers of embryos, from fertilization onward, can readily be obtained. Development is rapid and the large size of the embryos facilitates their micromanipulation. This allows analysis (at the single cell level if necessary) of such important issues as maternal regulation and embryonic inductions that are not feasible in mammalian, avian, or fish embryos. The course is suited both for those who have a knowledge of molecular biology and developmental biology but have had no experience with Xenopus, as well as those with some Xenopus experience who wish to learn newly developed, advanced techniques. The specific areas that will be covered are (1) Care and handling of Xenopus adults, removal of oocytes, induction of ovulation, egg collection, testis isolation, and in vitro and natural fertilization; (2) Stages of embryonic development and anatomy; (3) Whole mount in situ hybridization and immunohistochemistry, including double probe staining; (4) Microinjection of eggs and oocytes with lineage tracers, antisense morpholino oligonucleotides, mRNA, and DNA constructions; (5) Micromanipulation of embryos, including induction and transplantation assays; (6) Oligonucleotide antisense techniques for "knock-out" of maternal molecules; (7) Preparation and use of cell cycle extracts for construction of transgenic embryos; and (8) Use of Xenopus tropicalis as a genetic model. It is anticipated that the basic organization of the course and much of the course material will remain unchanged during the proposed period of support. However, fast-developing topics, such as the in vivo use of siRNA for gene knock-downs, new promoter sequences for use in transgenesis, and the availability of transgenic and mutant lines, will be discussed and integrated into the course as the field progresses.