农杆菌介导的针刺法将双价抗虫基因导入水稻

摘要

The plasmid of pCDMARUBA-Hyg containing two exogenous insect-resistant genes ( sbk and sck) was transformed into Agrobacterium EHA105, and the seed apical meristems of three rice varieties ( indica rice 9311, japonica rice Nanjing 45 and 110183) were directly infected by the above strain EHA 105 through needling inoculation .It was found that 10 plants of 9311, 5 plants of Nanjing 45, and 1 plant of 110183 seedlings survived .The results of conventional PCR detection showed that 9 plants of 9311, 3 plants of Nanjing 45, and 1 plant of 110183 seedlings were positive , and the total positive plants accounted for 81%of the total survived plants .After seminal root and bud were removed from the germinative seeds of Nanjing 45, its naked apical meristems were infected through needling inoculation , and 9 plants survived.PCR detection results indicated that: among the survived 9 plants, 4 plants contained divalent insect-resistant genes, 2 plants only contained gene sbk, and another 2 plants only contained gene sck.In comparison with the traditional method of Agrobacterium-mediated transformation , the present method does not need callus inducement and tissue culture , so it can save a large number of labors and costs .It is a simple, quick, efficient method for the transformation of rice genes .%用含2个抗虫基因的转化质粒pCDMARUBA－Hyg的农杆菌直接针刺侵染籼稻9311和粳稻南粳45、110183的种子胚芽生长点，对存活成苗的10株9311、5株南粳45和1株110183进行常规PCR检测，阳性株分别为9株、3株和1株，混合计算，阳性率为81％。去除南粳45发芽种子的种根和幼芽后，对裸露的胚芽生长点进行针刺接种，9株存活。经对成苗的PCR检测，4株含有2个外源基因，各有2株只含有1个基因（ sbk或sck）。与传统的农杆菌介导法相比，本研究的方法不需要诱导愈伤组织以及之后的一系列组织培养，基因转化时间缩短，节省了大量的人力和财力，具有简单、快速、高效之优点，是改良的水稻转基因新技术。