MDMV外壳蛋白基因的克隆及其在大肠杆菌中的表达

报道了玉米矮花叶病毒(MDMV)全长外壳蛋白cDNA基因的克隆、序列分析和表达产物鉴定的结果。根据其cDNA序列推测的外壳蛋白氨基酸顺序全长共有313个氨基酸,分子量总和为35 400 D,与聚丙烯酰胺凝胶电泳估算的值36 000 D相近,克隆的病毒外壳蛋白基因cDNA 3′-末端非编码区含有256个碱基。将此外壳蛋白的基因插入pUC19的1acZ基因中,转入E.coli后诱导表达,聚丙烯酰胺凝胶电泳分析呈现一特异性的蛋白带,经Western吸印与抗MDMV的兔血清呈阳性反应,证实所克隆的cDNA基因为MDMV外壳蛋白基因,并且能够正确表达。     

Introducing Bt Gene Into Maize With Ovary Injection

It is reported here that Bt toxin gene has been successfully transferred into maize inbred line by ovary injection for the first time both at home and abroad. One transgenic plant (To) has been confirmed by Southern blotting and PCR test, and 71 progenies (T1) from T0 have been obtained through self-pollination. Of these 71 progenies, seven plants demonstrated positive results in the PCR test; four were used to feed Asian corn borer, and certain effect of insect-resistance was observed. The experiments on the ovary injection in Hainan Province have also been repeated, thus providing new chance to the application of genetic engineering to the maize improvement.     

用于转化玉米的增效载体的鉴定及PEG介导的转化方法研究

本文用类质粒S-1构建了专门用于转化玉米的遗传工程载体pBIS5,通过PEG介导的转化方法,对玉米自交系BMS细胞的原生质体进行转化,转化频率比对照提高一倍,具有明显的增效作用。对PEG介导的转化方法进行了研究,发现PEG的浓度以6—10％(W/V)较好;PEG处理时间以30 min为最好,而DNA浓度则以40μg/ml为最好。上述参数配合特定的实验系统可以获得令人满意的转化效果。     

用子房注射法将Bt毒蛋白基因导入玉米的研究

本文在国内外首次报道了用子房注射的方法将Bt毒蛋白基因导入玉米自交系的全过程。获得的一株转基因玉米(T_0)自交留种,得到了71株T_1代的植株。转基因玉米(T_0)经点渍法、Southern吸印/分子杂交以及PCR扩增均获得阳性结果;用T_1代植株叶片DNA进行PCR扩增,在71株中有7株呈阳性反应;对其中4株进行抗玉米螟测试,呈现一定的抗虫效果;在海南岛用农大60玉米再次进行子房注射,也获得了成功,说明这种方法是可重复的,从而为利用基因工程技术改造玉米开拓了新的途径。