棉花法呢基焦磷酸合酶cDNA克隆、序列分析及其在种子发育过程中的表达特征

CLONING AND SEQUENCING OF A cDNA ENCODING FARNE SYL PYROPHOSPHATE SYNTHASE FROM GOSSYPIUM ARBOREUM AND ITS EXPRESSION PATTERN IN THE DEVELOPING SEEDS OF GOSSYPIUM HIRSUTUM cv. “SUMIAN 6” 

根据法呢基焦磷酸合酶（ＦＰＳ）保守域设计简并引物，应用ＮｅｓｔＰＣＲ和ＰＣＲ９６孔板筛库技术分离到亚洲棉（ＧｏｓｙｐｉｕｍａｒｂｏｒｅｕｍＬ．）法呢基焦磷酸合成酶的ｃＤＮＡ。序列分析表明，该ｃＤＮＡ全长１２８０ｂｐ，开放阅读框共编码３４２个氨基酸残基，推断蛋白质分子量约为３９ｋＤ，推测的氨基酸顺序与拟南芥和青蒿的ＦＰＰ合酶序列同源性为７８９％和８０７％。应用ＲＴＰＣＲ定量分析ｆｐｓ１基因在“苏棉６号”（Ｇ．ｈｉｒｓｕｔｕｍＬ．ｃｖ．“Ｓｕｍｉａｎ６”）种子发育过程中的表达特征，发现从胚珠开始形成种子（开花后２０ｄ）至种子成熟（开花后４０ｄ），ｆｐｓ１ｍＲＮＡ转录水平发生明显变化。在种子发育早期，开花后２０ｄ至２７ｄｆｐｓ１基因保持相对稳定的表达水平；但从开花后２７ｄ到种子成熟，ｆｐｓ１ｍＲＮＡ的转录水平迅速增高，此期倍半萜棉毒素在种子中的积累亦呈明显的增长趋势，完全风干成熟的种子中倍半萜棉毒素的含量可达１３ｍｇ／ｇ。推测ｆｐｓ１基因在转录水平参与种子中倍半萜类物质合成的调控。

A cDNA encoding farnesyl pyrophosphate (FPP) synthase was isolated from Gossypium arboreum L. cDNA library by using nest PCR and PCR 96 well plate screening methods. Nucleotide sequencing revealed that it is a full length cDNA of 1.28 kb, and the deduced peptide contained 342 amino acids, the putative amino acid sequence exhibited 80.7%, 78.9% and 71.6% identities with the FPP synthases of Artemisia annua, Arabidopsis thaliana and Zea mays, respectively. Quantitative RT PCR analyzed the steady state level of fps1 mRNA in the developing seeds of G.hirsutum L.cv. “Sumian 6”, the results revealed that the mRNA level was relatively stable in the early stages of the developing seeds, however, from 27 days post anthesis (DPA) to 40 DPA, transcription of fps1 mRNA was profoundly upregulated, the increment of the fps1 mRNA relative content was in combination with a concomitant accumulation of sesquiterpene aldehydes in the developing seed at this stage.