TA29-Barnase基因导致抗虫转基因欧洲黑杨雄性不育的研究  

A STUDY ON THE INTRODUCTION OF MALE STERILITY OF ANTIINSECT TRANSGENIC Populus nigra BY TA29 BARNASE GENE

作　　者：李玲[1] 齐力旺[1] 韩一凡[1] 汪迎春[2] 李文彬[2] 

机构地区：[1]中国林业科学研究院林业研究所,北京100091 [2]中国科学院遗传研究所,北京100101

出　　处：《林业科学》2000年第1期28-32,共5页Scientia Silvae Sinicae

基　　金：国家自然科学基金

摘　　要：取在１／２ＭＳ＋ ＮＡＡ０－０２ ｍｇ／Ｌ 培养基中发育４ ～６ 周的欧洲黑杨转Ｂｔ 基因植株１５ｎ１９２ 试管苗幼叶，在ＭＳ＋ ＢＡ０－２ ｍｇ／Ｌ＋ ＮＡＡ０－０２ ｍｇ／Ｌ培养基中培养１６ｈ ，分别用基因枪法和叶盘法转化ＴＡ２９Ｂａｒｎａｓｅ嵌合基因，其Ｂａｒｎａｓｅ 基因的转化率依次为１６－１ ％ 和２３ ％ ，再生株数分别为１７５／１２ 和７１／１０ 。试验表明，选择发育适宜的叶片，利于目的基因转化后的植株再生。进一步用ＳＤＳ 法提取转基因植株总ＤＮＡ，经ＰＣＲ 检测表明：转化植株基因组中扩增出的ＤＮＡ 片段与Ｂａｒｎａｓｅ 基因的大小一致；Ｓｏｕｔｈｅｒｎ Ｂｌｏｔ 分析出转化后的欧洲黑杨基因组中携有Ｂａｒｎａｓｅ 基因片段，进行Ｄｏｔ Ｂｌｏｔ 检测，转化植株有一明显的杂交斑，并证实为阳性转化植株，并通过试管培养，结合林木常规育种方法，获得了转Ｂａｒｎａｓｅ 基因的抗虫欧洲黑杨转基因植株。Chimaeric TA29 Barnase gene was introduced into anti insect transgenic Populus nigra of good quality and high yield by Agrobacterium tumefaciens(A.t)and biolistics transformation from tender leaves of 15 n 192 tube plantlets, which growing at the medium 1/2MS+NAA0 2?mg/L for 4～6 weeks, the leaves was cultured with MS+BA0 2?mg/L+NAA0 02?mg/L about 16 hours. The transformation frequency of Barnase gene by biolistics and A.t was 16 1 percent and 23 percent respectively, the number of repropagation plantlets were 175/12 and 71/10 respectively. The experiment showed that, the suitable developing leaves were selected, which was good for plantlets regeneration after Barnase gene transformation. Further more, we extracted the total DNA from transgenic plants by SDS method. The PCR analysis showed that, the DNA fragment from trangenic genome was amplified were completely identical with the Barnase gene; Southern Blot analysis showed that the transgenic plants genome carry about with Barnase gene fragment; Dot Blot analysis showed that the transgenic plants passes a cross spot clearly, and it was positive transformed plants further. The trans Barnasegenic plants of anti insect \%Populus nigra \%were obtained by the tissue culture and the methods of normal forest tree breeding.

关 键 词：抗虫转基因 欧洲黑杨 Barnase基因 雄性不育 

分 类 号：S792.110.4[农业科学—林木遗传育种]