Efficient Agrobacterium-mediated transformation of Lotus japonicus with reliable antibiotic selection
Efficient Agrobacterium-mediated transformation of Lotus japonicus with reliable antibiotic selection
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DOI：
10.1007/s00299-002-0494-4
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发表时间：
2002-10
影响因子：
6.2
通讯作者：
T. Aoki;A. Kamizawa;S. Ayabe
中科院分区：
文献类型：
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作者：
T. Aoki;A. Kamizawa;S. Ayabe
我们开发了一种新的转化模式豆科植物莲子叶的新方法，该方法从横子叶切段转化植株，而不会受到抗生素选择后存活的非转化子的污染。将HPT基因作为选择标记和GUS报告基因转化莲花叶，两者都由花椰菜花叶病毒35S启动子驱动。通过对转化子中GUS活性的测定，验证了潮霉素筛选的效果。基因组DNA的聚合酶链式反应分析也证实了GUS基因的整合。整合后的T-DNA稳定，为显性遗传性状。该方法在插入突变或基因标记的大规模转化中可能具有潜在的有效性和应用价值。
We have developed a new procedure for transforming a model legume,Lotus japonicus, that yields transformed plants from transverse cotyledon segments without contamination from the presence of non-transformants that survived the antibiotic selection.L. japonicuswas transformed with theHPTgene as a selectable marker and theGUSreporter gene, both of which were driven by cauliflower mosaic virus 35S promoter. The efficacy of selection with hygromycin was tested using the assay of GUS activity in putative transformants. The integration of theGUSgene was also confirmed by polymerase chain reaction analysis of the genomic DNA. The integrated T-DNA was stable and inherited as a dominant trait. This procedure may have potential effectiveness and application in large-scale transformation for insertional mutagenesis or gene tagging.