国家自然科学基金资助项目(30470156), 中国科学院创新项目(KZCX3-SW-215).
This work was supported by grants from The National Natural Science Foundation of China (30470156) and Key Innovative Project of Chinese Academy of Sciences (KZCX3-SW-215).
雨生红球藻是一种单细胞绿藻,在多种逆境胁迫条件下能够大量合成并迅速积累虾青素,其积累量最高可达细胞干重的4%,从而成为目前最理想的天然虾青素合成工具. 八氢番茄红素合成酶 (PSY) 是虾青素合成途径中第一个限速酶. 分离了八氢番茄红素合成酶基因 (psy) 的全长cDNA及基因组DNA. 其全长cDNA包括1 200个碱基,编码400个氨基酸,基因组DNA包括5个外显子,4个内含子. 系统发育分析结果显示,绿藻的八氢番茄红素合成酶基因形成一个进化枝,它们与高等植物的psy亲缘关系比较近. 通过Genome Walking的方法,分离了psy基因约1 kb的5′侧翼序列. 将含有TATA-box 和CAAT-box的297 bp的序列与LacZ报告基因构成嵌合的表达载体,用基因枪法转化雨生红球藻. lacZ的瞬间表达检测结果表明,这段上游序列能够驱动lacZ表达,具有启动子活性.
The unicellular green alga Haematococcus pluvialis accumulates a highly valuable ketocarotenoid, i.e. astaxanthin up to 4% dry weight under stress conditions. Phytoene synthase is considered to be the first rate limiting enzyme in carotenoid biosynthesis pathway in H. pluvialis. The cDNA and genomic genes of phytoene synthase, i.e. psy from H.pluvialis were cloned and characterized. Result showed that psy had one open reading frame of 1 200 bp encoding a putative polypeptide of 400 amino acids which was interrupted by four introns. Phylogenetic analysis revealed that psy from green algae formed a monophyletic clade, and its closer relationship was higher plants. By using genomic walking approach, an approximate 1 kb 5′ flanking region of psy gene was cloned and a number of putative cis-regulatory elements were revealed. Fusing a 297 bp internal sequence (-297 to -1 bp from the translation initiation codon of psy) with the reporter gene, i.e. lacZ before attempted introducing the construct into the green alga via particle bombardment resulted in lacZ transient expression.
梁成伟,赵方庆,秦 松,檀琮萍,魏 炜,孟春晓.雨生红球藻八氢番茄红素合成酶基因的克隆及表征[J].生物化学与生物物理进展,2006,33(9):854-860
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