Insertion in the coding region of the movement protein improves stability of the plasmid encoding a tomato mosaic virus-based expression vector
Insertion in the coding region of the movement protein improves stability of the plasmid encoding a tomato mosaic virus-based expression vector
复制标题
运动蛋白编码区的插入提高了编码基于番茄花叶病毒的表达载体的质粒的稳定性
DOI：
10.1007/s00705-008-0165-z
复制
发表时间：
2008
影响因子：
2.7
通讯作者：
M. Mori
中科院分区：
文献类型：
--
作者：
K. Dohi;A. Tamai;M. Mori
番茄花叶病毒（ToMV）遗传操作的主要障碍是含有ToMV载体的感染性全长cDNA的质粒的不稳定性，这常常阻止感兴趣的外源基因亚克隆到载体中。我们发现，在运动蛋白（MP）编码区插入0.3-1.6-kbp的DNA片段有效地减弱了质粒的细菌毒性，并大大提高了质粒产量。在MP编码区中含有0.3 kb插入的修饰的ToMV的积累与在烟草BY-2原生质体中没有插入的修饰的ToMV的积累相当，而超过0.6 kb的插入显著降低了病毒RNA的积累。将含有0.3kb插入片段的ToMV载体容易地操作以引入人干扰素-γ（HuIFN-γ）的编码序列，并成功地用于在BY-2原生质体和转基因BY-2细胞中产生HuIFN-γ。
A major obstacle in the genetic manipulation of tomato mosaic virus (ToMV) is the instability of the plasmid containing the infectious full-length cDNA of the ToMV vector, which often prevents the subcloning of a foreign gene of interest into the vector. We found that an insertion of a 0.3–1.6-kbp DNA fragment in the movement protein (MP) coding region effectively attenuated bacterial toxicity of the plasmid and greatly increased plasmid yield. Accumulation of a modified ToMV containing a 0.3-kb insertion in the MP coding region was comparable to that of a modified ToMV without an insertion in tobacco BY-2 protoplasts, while an insertion more than 0.6 kb significantly reduced accumulation of the viral RNA. The modified ToMV vector containing a 0.3-kb insertion was easily manipulated to introduce a coding sequence for human interferon-gamma (HuIFN-γ) and successfully utilized to produce HuIFN-γ in both BY-2 protoplasts and transgenic BY-2 cells.
影响因子：
5.4
作者：
Nishikiori, Masaki;Dohi, Koji;Ishikawa, Masayuki
通讯作者：
Ishikawa, Masayuki