Xiang C, Han P, Lutziger I, Wang K, Oliver D J
Department of Botany, Iowa State University, Ames 50011, USA.
Plant Mol Biol. 1999 Jul;40(4):711-7. doi: 10.1023/a:1006201910593.
A streamlined mini binary vector was constructed that is less than 1/2 the size of the pBIN19 backbone (3.5 kb). This was accomplished by eliminating over 5 kb of non-T-DNA sequences from the pBIN19 vector. The vector still retains all the essential elements required for a binary vector. These include a RK2 replication origin, the nptIII gene conferring kanamycin resistance in bacteria, both the right and left T-DNA borders, and a multiple cloning site (MCS) in between the T-DNA borders to facilitate cloning. Due to the reduced size, more unique restriction sites are available in the MCS, thus allowing more versatile cloning. Since the traF region was not included, it is not possible to mobilize this binary vector into Agrobacterium by triparental mating. This problem can be easily resolved by direct transformation. The mini binary vector has been demonstrated to successfully transform Arabidopsis plants. Based on this mini binary vector, a series of binary vectors were constructed for plant transformation.
构建了一种简化的微型双元载体,其大小不到pBIN19骨架(3.5 kb)的1/2。这是通过从pBIN19载体中去除超过5 kb的非T-DNA序列来实现的。该载体仍然保留了双元载体所需的所有基本元件。这些元件包括RK2复制起点、赋予细菌卡那霉素抗性的nptIII基因、左右T-DNA边界,以及位于T-DNA边界之间的多克隆位点(MCS)以方便克隆。由于载体尺寸减小,MCS中有更多独特的限制性酶切位点,从而允许进行更多样化的克隆。由于不包含traF区域,无法通过三亲本杂交将此双元载体导入农杆菌。这个问题可以通过直接转化轻松解决。已证明该微型双元载体能成功转化拟南芥植株。基于此微型双元载体,构建了一系列用于植物转化的双元载体。