Isopentenyl transferase gene expression offers the positive selection of marker-free transgenic plant of Kalanchoe blossfeldiana
Isopentenyl transferase gene expression offers the positive selection of marker-free transgenic plant of Kalanchoe blossfeldiana
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DOI：
10.1007/s11240-009-9519-9
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发表时间：
2009-03
期刊：
影响因子：
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通讯作者：
G. Thirukkumaran;R. Khan;D. Chin;I. Nakamura;M. Mii
中科院分区：
文献类型：
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作者：
G. Thirukkumaran;R. Khan;D. Chin;I. Nakamura;M. Mii
允许生产没有选择标记基因的转基因植物的技术在公共和环境安全方面具有极大的兴趣。为了产生这种无标记的转基因植物，多自动转化[MAT]载体系统提供了可能性，该系统将使用异戊烯基转移酶（ipt）基因的阳性选择与产生无标记植物的位点特异性重组相结合。本研究以根癌农杆菌EHA 105为出发菌株，构建了含有lacZ、gus基因和T-DNA区可去除盒的pMAT 21载体，以ipt基因为选择标记基因。共培养的外植体培养在无激素和选择剂的MS培养基上，85%的再生芽表现出分化芽表型，GUS表达。在继代培养过程中产生了41个形态正常的芽。通过PCR分析确定，超过90%的正常芽为ipt −，gus−butlacZ+。这些结果表明，ipt表型与非转基因以及转基因无标记芽明显不同。本研究为无标记转基因的产生开辟了有趣的前景。目的有用的转基因拟南芥。
The technologies allowing the production of transgenic plants without selectable marker genes, is of great interest in public and environmental safety. For generating such marker-free transgenic plants, possibility has been offered by Multi-Auto-Transformation [MAT] vector system, which combines positive selection, using the isopentenyl transferase (ipt) gene, with a site-specific recombination that generates marker-free plants. In this studyAgrobacterium tumefaciensstrain EHA105 harboring anipt-type MAT vector,pMAT21, containinglacZ, gusgenes and the removable cassette in the T-DNA region was used to produce marker-free transgenicKalanchoe blossfeldianaPoelln., employingiptgene as the selectable marker gene. Co-cultivated explants were cultured on hormone- and selective agent-free MS medium, and 85% of the regenerated shoots showedipt-shooty phenotype with GUS expression. Forty-one morphologically normal shoots were produced during the subculture. More than ninety percent of the normal shoots wereipt−,gus−butlacZ+as determined by PCR analyses. These results indicate that theiptphenotype was clearly distinguishable from non-transgenic as well as transgenic marker-free shoots. This study opens interesting perspective for the generation of marker-free transgenicK. blossfeldianawith objective useful transgene.