Isolation and analysis of mutants with altered ABA recognition specificity using an ABA antagonist.

为了鉴定阿坝受体，我们采用了一种新的筛选方法，使用阿坝结合抑制剂分离出具有改变的阿坝受体结合特异性的突变体。我们使用阿坝类似物PBI-51作为阿坝结合抑制剂，因为该化学物质已被报道在甘蓝型油菜中作为阿坝结合抑制剂起作用。我们尝试用两种筛选策略分离突变体。一个是寻找似乎识别PB 151作为阿坝结合抑制剂的突变体。我们从EMS处理的植物的500，000粒M2种子中分离出几种候选物。然而，这些候选人原来是阿坝不敏感的突变体。另一种策略是分离似乎将PB 151识别为阿坝的突变体。在该筛选中，我们从EMS处理的植物的400，000个M2种子中分离出8个独立的品系。这些候选突变体与已知的ABA超敏突变体eal和ein 2不是等位基因，但都表现出ABA超敏性。为了提高阿坝受体突变体的分离效率，我们提出了利用aba 2突变体的策略，并分离了几十个候选突变体。这些突变基因的鉴定将有助于我们对阿坝信号转导途径的深入了解。在分离阿坝相关突变体的过程中，我们从拟南芥中分离到一个新的乙醇敏感突变体gekol。我们通过定位鉴定了GEKO I基因，发现它编码一种新的蛋白质，在植物和古细菌中是保守的。GEKO I可能在植物特异性功能中起关键作用。

In order to identify the ABA receptors, we employed a novel screening using an ABA binding inhibitor to isolate mutants that have altered binding specificity of ABA receptors. We used a ABA analogue, PBI-51, as a ABA binding inhibitor since this chemical has been reported to work as ABA binding inhibitor in Brassica napus. We tried to isolate mutants with two type of screening strategies. One is to search for mutants that seem to recognize PB151 as an ABA binding inhibitor. We isolated several candidates from 500,000 M2 seeds from EMS treated plants. However, these candidates turned out to be ABA insensitive mutants. The other strategy was to isolate mutants that seems recognize PB151 as ABA. In this screen, we isolated 8 independent lines from 400,000 M2 seeds from EMS treated plants. Those candidates show ABA-hypersensitivity, although these are not allelic to known ABA-hypersensitive mutants, eral or ein2.To increase the efficiency to isolate ABA receptor mutants, we thought out the strategy to use aba2 mutants, and isolated several dozens of candidates. ldentification of those mutated genes will surely contribute in our understanding on ABA siganling pathway. In the course to isolate ABA related mutants, we isolated a novel ethanol hypersensitiv mutant, gekol, from Arabidopsis. We identified the GEKO I gene by mapping and found it encoded a novel protein that conserved among plants and archea bacteria. GEKO I may play a pivotal role for plant sp ecific functions.