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An approach to the biochemical mechanisms of heartwood formation using cell cultures

利用细胞培养研究心材形成的生化机制的方法

基本信息

批准号：
03454081
负责人：
SAKAI Kokki
金额：
$ 4.16万
依托单位：
KYUDHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1991
资助国家：
日本
起止时间：
1991 至 1993
项目状态：
已结题

项目摘要

(1). Biochimistry of lignificationLignification process has been investigated in this project since it occurs prior to the heartwood formation in forest tree cells. A peroxidase isoenzyme was found to catalyze substrate specifically the dehydrogenetive polymerization of sinapyl alcohol. This enzyme was isolated and characterized from cell cultures of Populus alba.(2). Establishment of cell cultures that produce heartwood constituentsCallus cultures of Cupressus lusitanica (Cupressanceae) and Pinus spp. produced their heartwood constituents, beta-thujaplicin and pinosylvins, respectively, although no heartwood constituents were detected in calluses of Cryptomeria japonica and Chamaecyparis obtusa. Thus, the culture of C.lusitanica cells is regarded as a good experimental material for studying phenomena of heartwood formation.(3). Elicitor of beta-thujaplicin accumulation in callus culturesThe addition of yeast extract to callus cultures of C.lusitanica leads to a large increase in the p … More roduction of beta-thujaplicin that act as a phyoalexin of the treated cultures. The extract was fractionated by means of ethanol precipitation, ribonucleasetreatment, gel permiation choromatography and Cu-complexation. The most effective fraction was a polysaccharide composed of maily mannose and glucose.(4). Biosythetic pathway of beta-thujaplicinCallus cultures of C.lusitanica was fed with ^<14>C-mevaronate, ^<14>C-glucose or ^<14>C-malonate by the shot-gun method. Elicitor was added to the callus at the same time. The results proved that beta-thujaplicins synthsized via the mevaronate pathway.(5). An attempt to isolation of the key enzyme(s) for beta-thujaplicin biosynthesisWe tried to isolate any enzyme(s) that catalyze biosynthesis of beta-thujaplicin from isopentenyl pyrophosphate. However, such enzyme(s) have not been detected in cell-free extract of callus cultures of C.lusitanica. Isoltion of the key enzyme(s), its characterization and proof of its presence in transition zone of C.lustanica trees should be subjects to be solved in the near future. Less

(1).木质化的生物化学本项目对木质化过程进行了研究，因为它发生在林木细胞心材形成之前。发现过氧化物酶同工酶特异性催化芥子醇的脱氢聚合底物。该酶是从白杨细胞培养物中分离出来并进行表征的。(2)。建立产生心材成分的细胞培养物Cupressus lusitanica（柏科）和松属植物的愈伤组织培养物。尽管在柳杉和日本扁柏的愈伤组织中没有检测到心材成分，但分别产生了其心材成分 β-thujaplicin 和 pinosylvins。因此，C.lusitanica细胞培养物被认为是研究心材形成现象的良好实验材料。(3)．愈伤组织培养物中 β-thujaplicin 积累的引发剂在 C.lusitanica 愈伤组织培养物中添加酵母提取物，会导致 β-thujaplicin 的产量大幅增加，β-thujaplicin 充当处理后培养物的植物抗毒素。通过乙醇沉淀、核糖核酸酶处理、凝胶渗透色谱法和铜络合对提取物进行分级。最有效的部分是主要由甘露糖和葡萄糖组成的多糖。(4)。 β-thujaplicin的生物合成途径通过鸟枪法向C.lusitanica的愈伤组织培养物添加14 C-甲羟戊酸、14 C-葡萄糖或14 C-丙二酸。同时将诱导子添加到愈伤组织中。结果证明β-thujaplicins是通过甲羟戊酸途径合成的。(5)．分离β-thujaplicin生物合成关键酶的尝试我们尝试从异戊烯基焦磷酸中分离出催化β-thujaplicin生物合成的任何酶。然而，在 C.lusitanica 愈伤组织培养物的无细胞提取物中尚未检测到此类酶。关键酶的分离、其表征以及其在 C.lustanica 树过渡区中存在的证明应该是在不久的将来需要解决的课题。较少的