Gene-technological manipulation of phospholipid composition in Arabidopsis thaliana in relation to freezing tolerance

(1)综合可供订货量：磷酸乙醇胺胞苷酰转移酶（EC2.7.7.14，ECT）是植物细胞中磷脂酰乙醇胺生物合成的限速酶。我已经通过PCR克隆了拟南芥ECT的cDNA，使用从拟南芥基因组数据库（Accession no.g3786005）设计的引物。(2)通过植物体内转化法将拟南芥CTP：磷酸胆碱胞苷酰转移酶（EC2.7.7.15，CCT）cDNA文库导入拟南芥。其中一个转化体表现出磷脂水平的变化。脂成分的变化与抗冻性之间的相关性正在研究中。(3)利用构建的甘蓝型油菜根cDNA文库pFL 61（URA 3）转化INY 102 B/YCpGPSS。从1 x 10^6个Ura^+转化子中分离出100个Etn^+转化子，并对其中16个进行了鉴定。它们都是相同的，并且代表性克隆包含1，894 bp和1，473 bp的开放阅读框。推导的氨基酸序列含有490个氨基酸残基，计算分子量为55，153。一个同源的拟南芥cDNA克隆通过BLAST搜索拟南芥EST序列的特点。拟南芥克隆含有1，739 bp，开放阅读框为1，449 bp。推导的氨基酸序列包含482个氨基酸残基，计算分子量为54，153。在GenBank核苷酸序列数据库中搜索同源性显示，两个克隆均与吡哆醛-5 '-磷酸依赖性组氨酸脱羧酶同源，并且两种蛋白质中均保守有一个推定的II组脱羧酶的磷酸吡哆醛结合基序（Sandmeier et al. 1994）。因此，我们的研究结果是第一个建议，组胺可以在植物根部合成。

(1) CTP : phosphoethanolamine cytidylyltransferase (EC2.7.7.14, ECT) is a rate-limiting enzyme in the biosynthesis of phosphatidylethanolamine in plant cells. I have cloned a cDNA for Arabidopsis ECT by PCR, using primers designed from Arabidopsis genome databases (Accession no. g3786005).(2) A cDNA for Brassicanapus CTP : phosphocholine cytidylyltransferase (EC2.7.7.15, CCT) library was introduced into Arabidopsis thaliana by an inplanta transformation method. One of transformants showed changes in the levels of phospholipids. Correlation between the changes in the lipid composition and frost resistance is under investigation.(3) INY102B/YCpGPSS was transformed with the pFL61 (URA3) cDNA library constructed from roots of Brassica napus. From 1 x 10^6 Ura^+ transformants, 100 Etn^+ transformants were isolated, and 16 of them were characterized. They are all identical and a representative clone contained 1,894 bp with an open-reading frame of 1,473 bp. Deduced amino-acid sequence contained 490 amino-acid residues with a calculated molecular mass of 55,153. A homologous Arabidopsis cDNA clone identified by a BLAST search to Arabidopsis EST sequences was also characterized. The Arabidopsis clone contained 1,739 bp with an open-reading frame of 1,449 bp. Deduced amino-acid sequence contained 482 amino-acid residues with a calculated molecular mass of 54,153. Search for homology in GenBank nucleotide sequence database revealed that both clones are homologous to pyridoxal-5'-phosphate-dependent histidine decarboxylases, and a putative pyridoxal phosphate-binding motif for Group II decarboxylases (Sandmeier et al. 1994) was conserved in both proteins. Therefore, our results are the first to suggest that histamine could be synthesized in plant roots.