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以奥林达夏橙[Citrus sinensis(L.)‘Olinda’]为材料,克隆了CsTBL1基因编码起始位点上游长度为2 361 bp的启动子序列。生物信息学预测表明,该启动子中含有多个与逆境应答相关的顺式作用元件。为进一步分析CsTBL1启动子的功能,构建了该基因启动子与GUS基因融合的植物表达载体,并用浸花法转化拟南芥,对T3代转基因拟南芥进行GUS活性染色。结果显示,CsTBL1启动子在1～3 d龄幼苗所有组织中的活性都非常强,在7～20 d龄幼苗的子叶和根中的活性仍然较强,但在下胚轴中基本检测不到活性。在50 d龄幼苗中,只在叶、茎的毛状体以及还在生长的根中检测到GUS活性。转基因植株的GUS表达受到1–氨基环丙烷–1–羧酸(乙烯合成前体,ACC)、脱落酸(ABA)、甲基茉莉酸(MeJA)和水杨酸(SA)诱导。上述结果表明CsTBL1可能在植物发育和抗外界逆境胁迫中起到非常重要的作用。
The promoter sequence of 2 361 bp upstream of CsTBL1 gene was cloned from Citrus sinensis (L.) Olinda. Bioinformatics prediction shows that the promoter contains multiple cis-acting elements related to stress response. To further analyze the function of CsTBL1 promoter, a plant expression vector fused with GUS gene promoter was constructed. Arabidopsis thaliana was transformed by immersion method and GUS activity was stained for T3 generation transgenic Arabidopsis thaliana. The results showed that the activity of CsTBL1 promoter was very strong in all tissues of 1-3 days old seedlings, and its activities in cotyledons and roots of seedlings at 7-20 days old were still strong. However, little activity was detected in hypocotyls . In 50-day-old seedlings, GUS activity was detected only in the leaf trichomes as well as in the growing roots. GUS expression in transgenic plants was induced by 1-aminocyclopropane-1-carboxylic acid (ethylene synthesis precursor, ACC), abscisic acid (ABA), methyl jasmonate (MeJA) and salicylic acid (SA). The above results indicate that CsTBL1 may play a very important role in plant development and resistance to environmental stress.