抱茎独行菜MUM4基因的克隆及分析(英文)  

Cloning and Analysis of Seed Coat Mucilage-related Gene MUM4 fromLepidium perfoliatum

作　　者：袁军文[1,2] 黄代红[1] 徐栋生[1] 赵娟[1] 李玲[1] 兰海燕[1] 张富春[1] 

机构地区：[1]新疆大学生命科学与技术学院新疆生物资源基因工程重点实验室,乌鲁木齐830046 [2]新疆生产建设兵团种子管理总站,乌鲁木齐830011

出　　处：《西北植物学报》2013年第10期1940-1952,共13页Acta Botanica Boreali-Occidentalia Sinica

基　　金：国家自然科学基金(30860020);中国教育部留学回国学者基金项目(2006-2008)

摘　　要：抱茎独行菜(Lepidium perfoliatum L.)为十字花科具典型粘液繁殖体植物,为探究该植物中种皮粘液质基因(MUCILAGE-MODIFIED4,MUM4,该基因在拟南芥中编码NDP-L-鼠李糖合成酶)的功能,通过生物信息学分析设计引物克隆得到抱茎独行菜MUM4基因,命名为LpMUM4。同源比对分析结果表明,LpMUM4与拟南芥AtMUM4基因具有很高的一致性。qRT-PCR结果表明,该基因在抱茎独行菜各组织中均有表达,在角果和根中的表达量最高,且其表达量随角果的发育表现出渐强的趋势。免疫组织化学定位分析表明,LpMUM4基因于角果发育的早期阶段在内珠被和外珠被都有表达,而在外珠被的表皮和亚表皮中表达量更高,至角果发育的最后阶段,其表达集中于表皮和亚表皮层,这可能与抱茎独行菜的外珠被发育成种皮及粘液质的生成有关。将LpMUM4基因转化拟南芥,该基因的过表达对位于粘液质合成途径中的上游基因AtTTG1具有显著的抑制作用。表型比对观察显示,转基因拟南芥与其野生型植株形态无显著差异,这可能是因为抱茎独行菜种皮的发育和粘液质的形成是一个多基因调控的复杂过程,某一基因的过表达或许不会引起明显的表型变化。During differentiation,the Lepidium perfoliatum seed coat epidermal cells produce copious amounts of mucilage that is extruded from the seed coat upon imbibition.Mucilage is composed primarily of pectin,a polysaccharide that is a main component of the cell wall.To validate the function of a mucilage related gene MUM4 （MUCILAGE-MODIFIED4,encodes a putative NDP-l-rhamnose synthase in Arabidopsis）,LpMUM4 from L.perfoliatum,which is myxospermous,was characterized in the present study.Sequence analysis revealed that the protein is highly homologous to AtMUM4 from Arabidopsis.Analysis of qRT-PCR showed that LpMUM4 was expressed in various parts of L.perfoliatum,and the transcription level was relatively high in silicles and root and the expression level gradually increased with the development of silicles.Moreover,the immunolocalization analysis showed that LpMUM4 was strongly expressed in both outer and inner integument layers at the earlier stage,then was observed more strongly in the epidermal and subepidermal layers of outer integument,finally was concentrated in between the epidermal and subepidermal layers,which corresponded to the process of the outer integument （testa） differentiation and the mucilage production of seed in L.perfoliatum.Overexpression of LpMUM4 in Arabidopsis showed a remarkable inhibition in the transcription level of AtTTG1 gene （which locates upstream of the AtMUM4 in the mucilage synthesis pathway） in silicles.However,the inspection of the morphology and mucilage release behavior of seed coat revealed no evident difference between LpMUM4 overexpressed transgenic line and WT.The possible explanation may be that the differentiation of the seed coat and production of the mucilage are complex processes in L.perfoliatum,and which is regulated by an interaction network of numerous genes with function redundancy.Therefore,the increasing expression of one gene （LpMUM4） during development may not cause evident phenotype change.

关 键 词：抱茎独行菜 粘液质合成 免疫定位 MUM4基因 QRT-PCR 转基因植物 

分 类 号：Q786[生物学—分子生物学]