Collaborative Research: Cell Walls Resistant to Environmental Extremes: the Chlamydomonas zygaspore as a Model System

孢粉素，通常被称为地球上最具抗性的生物聚合物，是成熟花粉粒的重要组成部分。孢粉素也存在于非种子植物的孢子壁和各种微生物的静息细胞中。尽管这种明显古老的化合物很重要，但孢子孢素的合成、分泌和组装途径仍不清楚。单衣藻（Chlamydomonas monoica）的合子壁含有孢粉素，这种单细胞藻类合子突变体的存在表明单衣藻的发育可以作为研究孢粉素生物合成和功能的模型。在野生型和突变型菌株中，通过荧光或透射电子显微镜和细胞壁形态发生过程中溶解的蛋白质的电泳，将利用差异固定和染色来分析合子细胞壁的组成。核转化程序将被开发，负责合子缺陷的基因的野生型等位基因将通过它们补充突变表型的能力被分离出来，使用一个索引的cosmid文库进行转化。或者，插入成熟突变体将通过核转化产生，然后分子标记将被用来克隆缺陷基因。通过这些方法克隆的基因的表达将在野生型和成熟突变体中进行比较。

9808478 Stern Sporopollenin, often referred to as the most resistant biopolymer on earth, is an essential component of mature pollen grains. Sporopollenin is also found in the spore walls of non seed plants, and in the resting cells of diverse microorganisms. Despite the importance of this apparently ancient compound, the routes of synthesis, secretion, and assembly of sporopollenin remain unknown. The zygospore wall of Chlamydomonas monoica contains sporopollenin and the availability of zygospore mutants of this unicellular alga suggests development of C. monoica as a model for the study of sporopollenin biosynthesis and function. The composition of the zygospore wall will be analyzed using differential fixation and staining followed by fluorescence or transmission electron microscopy and by electrophoresis of proteins solubilized during wall morphogenesis in wildtype and mutant strains. A nuclear transformation procedure will be developed and wildtype alleles of genes responsible for zygospore defects will be isolated by their ability to complement mutant phenotypes, using an indexed cosmid library for transformation. Alternatively, insertional maturation mutants will be generated by nuclear transformation and the molecular tag will then be used to clone the defective gene. The expression of genes cloned by these approaches will be compared in wildtype and maturation mutants.