Stable transformation ofArabidopsis with thebar gene using particle bombardment
Stable transformation ofArabidopsis with thebar gene using particle bombardment
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DOI：
10.1007/bf01973587
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发表时间：
1994-09
影响因子：
3
通讯作者：
T. Sawasaki;M. Seki;H. Anzai;K. Irifune;H. Morikawa
中科院分区：
文献类型：
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作者：
T. Sawasaki;M. Seki;H. Anzai;K. Irifune;H. Morikawa
构建了在花椰菜花叶病毒 (CaMV) 35S 启动子和胭脂碱合酶 (NOS) 终止子控制下编码草胺膦乙酰转移酶 (PAT) 的 bar 基因的质粒 pARK 22，并使用气动粒子枪将其引入拟南芥根切片中。用该质粒轰击的根切片产生的耐药愈伤组织的产量比用分别含有新霉素磷酸转移酶和潮霉素磷酸转移酶基因的pCaMVNEO或pCH轰击的根切片高四到八倍。在从独立的双丙氨磷抗性愈伤组织获得的许多初级转化体(T0)植物中，通过Southern印迹杂交和PAT酶活性分析对其中三种进行了研究，证实了外源基因稳定整合到拟南芥基因组中及其在植物中的表达。后代分析显示外源基因的传递及其表达直至T2代。一些T1后代表现出形态异常。因此，可以有效地使用bargene来选择转基因A。拟南芥植物。
A plasmid pARK 22 harbouring thebargene encoding phosphinothricin acetyltransferase (PAT) under the control of the cauliflower mosaic virus (CaMV) 35S promoter and nopaline synthase (NOS) terminator was constructed and introduced into root sections ofArabidopsis thalianausing the pneumatic particle gun. The root sections that had been bombarded with this plasmid gave four to eight times higher yield of drug-resistant calluses than those sections bombarded with pCaMVNEO or pCH, which respectively contain the neomycin phosphotransferase and hygromycin phosphotransferase genes. Among a number of primary transformant (T0) plants obtained from independent bialaphos-resistant calluses, three were studied by Southern blot hybridization and PAT enzyme activity analyses, confirming the stable integration of the foreign gene into theArabidopsisgenome and its expression in plants. The progeny analysis showed transmission of the foreign gene and its expression in up to the T2generation. Some of the T1progeny showed morphological abnormalities. Thus, thebargene can be used effectively to allow selection of transgenicA. thaliannaplants.