棉花种子特异表达的LEA启动子克隆及功能验证

Isolation and Functional Characterization of the Seed-specific Promoter of LEA Gene from Cotton(Gossypium hirsutum L.)

以棉花品种新陆早33号为材料,克隆获得其胚胎发育晚期丰富蛋白LEA基因的种子特异性启动子。启动子序列全长为1228bp;作用元件分析表明该区域除了具有启动子核心调控序列外,还含有多个与组织特异性、激素、逆境等表达相关的顺式作用元件,如E-box、ABRE元件、A-box等。与已报道的棉花品种Coker 201的LEA基因D34的5'端上游调控序列1212bp相比,两者具有97%的一致性。拟南芥遗传转化的功能分析结果表明,所克隆的序列能驱动GUS基因在种子中特异表达,且GUS主要在转基因植物的种子发育后期表达;其表达强度要弱于组成型的CaMV35S启动子。研究结果不仅有助于进一步深入认识棉花LEA基因功能及其表达调控规律,也为植物遗传转化提供组织特异性的启动子。

A seed-specific promoter of LEA（late embryogenesis abundant） protein gene was successfully cloned from cotton vari- ety Xinluzao 33（XLZ33） in this study. Its complete nucleotide sequence was 1228 bp. C/s-acting elements search in the segment revealed the presence of a number of motifs including E-box, ABRE motif and A-box, etc. The cloned sequence showed 97% i- dentity with a 1212 bp fragment, 5＇ upstream regulatory sequence of LEA gene D34, reported from cotton variety Coker 201. Functional analysis in transgenic Arabidopsis plants showed the cloned fragment could drive GUS reporter gene to express ex- clusively in seeds. GUS activity was detected mainly at the late stage of seed development of transgenic plants. The lower tran- scriptional levels of GUS were detected in transgenic plants containing LEA promoters than in transgenic plants with the consti- tutive CaMV35S promoter. The results may be helpful for our understanding of the spatial and temporal regulation mechanisms of the LEA gene, and also provide a tissue-specific promoter that could be applicable to the modification of seed phenotypes of transgenic plants.