Development of recycling biomass for recycling systems of organic waste

开发用于有机废物回收系统的回收生物质

基本信息

批准号：
15580297
负责人：
SUGIMOTO Manabu
金额：
$ 2.05万
依托单位：
Okayama University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2005
项目状态：
已结题

项目摘要

The genes encoding pro-form and active-form of strong fibrinolytic protease, isozme A, in the six isozymes of eartworm, Lumbricus rubellus, were subcloned into a plasmid, pEU3-NII, and expressed in the wheat germ cell-free protein synthesis system. Extra proteins with a molecular mass of about 26 and 25 kDa were produced by pro-form and active-form of plasmids, respectively, which are similar to calculated molecular mass from the amino acid sequences. These results shows that the gene from earthworm could express in plant.The pro-form and active-form genes of isozyme A were subcloned into a plasmid pBI121, resulting the plasmids both of which the enzyme genes fused with CaMV35S promoter. These genes were transferred to Arabidopsis thaliana, cv. Columbia, and T1 seeds were obtained. T1 seeds including active-form gene were cultured on MS medium agar plate containing kanamycine and three types of plants, 1) normal shape, 2) breached leaf, 3) wrinkled leaf, were observed. All types of transgenic plants expressed extra isozyme A mRNA, but could not produce T2 seeds. The transgenic plants of T1 seeds including pro-form gene also showed the wrinkled leaves which is breached gradually, resulting the die. These results suggest that the isozyme A from earthworm is produced and works in plants, and the possibility of producing the organic waste-degrading plants if the expression of the isozyme A gene could be controlled by adequate promoters which can restrict the expression strongly or organ-specific expression affecting little on plant growth.

在eart虫的六个同工酶中编码强纤维蛋白蛋白酶的促形式和活性形式的基因，将lumbricus rubellus的六个同工酶亚克隆到质粒中PEU3-NII，并在小麦生殖细胞细胞中表达。分子质量分子质量约为26和25 kDa的额外蛋白质分别通过质粒和活性形式产生，这些质粒与氨基酸序列的分子质量相似。这些结果表明，来自earth的基因可以在植物中表达。同工酶A的促形式和活性形式的基因被亚克隆到质粒PBI121中，从而导致质粒与CAMV35S启动子融合的酶基因。这些基因被转移到拟南芥，简历。获得哥伦比亚和T1种子。在含有卡纳霉素的MS中琼脂板上培养了包括活性形式基因的T1种子，三种类型的植物，1）正常形状，2）损坏的叶子，3）皱纹的叶片。所有类型的转基因植物都表示额外的同工酶A mRNA，但无法产生T2种子。 T1种子的转基因植物在内还显示了逐渐破裂的皱纹叶子，导致死亡。这些结果表明，从earth产生同工酶A并在植物中起作用，并且如果同工酶的表达可以通过适当的启动子来控制，那么产生有机废物降解的植物的可能性，可以强烈或有机体限制表达的表达。具体表达对植物生长的影响很小。