Frank M J, Liu D, Tsay Y F, Ustach C, Crawford N M
Department of Biology, University of California at San Diego, La Jolla 92093-0116, USA.
Plant Cell. 1997 Oct;9(10):1745-56. doi: 10.1105/tpc.9.10.1745.
Tag1 is a transposable element first identified as an insertion in the CHL1 gene of Arabidopsis. The chl1::Tag1 mutant originated from a plant (ecotype Landsberg erecta) that had been transformed with the maize transposon Activator (Ac), which is distantly related to Tag1. Genomic analysis of untransformed Landsberg erecta plants demonstrated that two identical Tag1 elements are present in the Landsberg erecta genome. To determine what provides transposase function for Tag1 transposition, we examined Tag1 excision in different genetic backgrounds. First, the chl1::Tag1 mutant was backcrossed to untransformed wild-type Arabidopsis plants to remove the Ac element(s) from the genome. F2 progeny that had no Ac elements but still retained Tag1 in the CHL1 gene were identified. Tag1 still excised in these Ac-minus progeny producing CHL1 revertants; therefore, Ac is not required for Tag1 excision. Next, Tag1 was inserted between a cauliflower mosaic virus 35S promoter and a beta-glucuronidase (GUS) marker gene and transformed into tobacco. Transformants showed blue-staining sectors indicative of Tag1 excision. Transgenic tobacco containing a defective Tag1 element, which was constructed in vitro by deleting an internal 1.4-kb EcoRI fragment, did not show blue-staining sectors. We conclude that Tag1 is an autonomous element capable of independent excision. The 35S-GUS::Tag1 construct was then introduced into Arabidopsis. Blue-staining sectors were found in cotyledons, leaves, and roots, showing that Tag1 undergoes somatic excision during vegetative development in its native host.
标签1是一种转座元件,最初被鉴定为插入拟南芥CHL1基因中。chl1::标签1突变体源自一株用玉米转座子激活子(Ac)转化的植物(生态型兰茨贝格直立型),Ac与标签1的亲缘关系较远。对未转化的兰茨贝格直立型植物的基因组分析表明,兰茨贝格直立型基因组中存在两个相同的标签1元件。为了确定是什么为标签1转座提供转座酶功能,我们在不同的遗传背景下检测了标签1的切除情况。首先,将chl1::标签1突变体与未转化的野生型拟南芥植物回交,以从基因组中去除Ac元件。鉴定出没有Ac元件但CHL1基因中仍保留标签1的F2后代。标签1在这些不含Ac的后代中仍然切除,产生CHL1回复体;因此,标签1切除不需要Ac。接下来,将标签1插入花椰菜花叶病毒35S启动子和β-葡萄糖醛酸酶(GUS)标记基因之间,并转化到烟草中。转化体显示出蓝色染色区域,表明标签1发生了切除。含有通过删除内部1.4 kb EcoRI片段体外构建的缺陷标签1元件的转基因烟草没有显示出蓝色染色区域。我们得出结论,标签1是一个能够独立切除的自主元件。然后将35S-GUS::标签1构建体导入拟南芥。在子叶、叶片和根中发现了蓝色染色区域,表明标签1在其天然宿主的营养发育过程中发生体细胞切除。