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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

（一）.木质化过程发生在林木细胞心材形成之前，因此本项目对木质化过程进行了研究。发现一种过氧化物酶同工酶能特异性催化芥子醇的聚合反应。该酶是从银白杨细胞培养物中分离和表征的。（二）、产生心材成分的细胞培养体系的建立墨西哥柏和松树的愈伤组织培养虽然在柳杉和钝头Chamaparis obtusa的愈伤组织中没有检测到心材成分，但它们分别产生了心材成分β-崖柏素和松素。因此，葡萄牙柳杉细胞培养是研究心材形成现象的良好实验材料。（三）、愈伤组织培养中β-崖柏素积累的诱导剂向墨西哥崖柏愈伤组织培养物中添加酵母提取物导致愈伤组织中β-崖柏素积累的大量增加。 ...更多信息 β-崖柏素的产生，所述β-崖柏素充当经处理的培养物的藻抗毒素。提取物通过乙醇沉淀、核糖核酸酶处理、凝胶渗透层析和铜络合等方法进行分级。最有效的组分是主要由甘露糖和葡萄糖组成的多糖。（四）、β-崖柏素的生物合成途径用鸟枪法分别用β-<14>C-甲羟萘酸、β-<14>C-葡萄糖和β<14>-C-丙二酸对墨西哥崖柏愈伤组织进行培养。同时向愈伤组织中加入激发子。结果表明，β-thujaplicins通过甲羟戊酸途径降解。（五）、β-崖柏素生物合成关键酶的分离我们试图从异戊烯焦磷酸中分离出催化β-崖柏素生物合成的酶。然而，这样的酶（S）没有检测到C. lusitanica的愈伤组织培养物的无细胞提取物。关键酶的分离、鉴定及其在过渡区的存在证明是今后亟待解决的问题。少