玉米基于Ac/Ds转座子的激活标签突变体生成系的构建

An artificial transposon Dissociation (Ds) element carries the cauliflower mosaic virus (CaMV) 35S enhancer will be inserted in maize Rsc gene. The construct based on skeleton vector pTF102 will be introduced into maize tissue culture by A.tumefaciens-mediated transformation. The positive plants will hybrid with maize inbreed T3301-56 which containing a activator transposase(Ac-Immobilized), then selfing and identify the homozygous plants. Then an activation-tagging Ac-Ds transposon system will be developed. After cultivation, purple grains from maize ear will be selected and yellow grains discarded. This Ac/Ds-based activation tagging system in maize(Zea mays. L) will be exploit to generate gain-of-function phenotype mutants for functional genomics research and for maize breeding.

将来源于CaMV35S启动子的增强子序列合成在玉米Ds转座子的左右两端重复序列中，这个包含有增强子序列的人工Ds转座子将插入在玉米Rsc基因之中，使Rsc失去活性。以上的所有DNA序列将构建在载体pTF102上。构建成功的转化质粒，由农杆菌介导转化玉米，得到的阳性转基因玉米材料与含有转座酶Ac的玉米自交系T3301-56杂交，在杂交后代中鉴定自交纯合的转基因材料，即为玉米基于Ac/Ds的激活标签突变体生成系。按照玉米生产的常规方法，大量种植激活标签突变体生成系，授粉自交结实。收集玉米果穗上大部分黄色籽粒中紫红色的籽粒，大量种植，观察植株的表型，自交保存，它们构成了玉米的激活标签突变体库。克隆分析插入标签的特异序列，总结该突变体生成系的规律，并从中筛选各种类型的功能获得型突变体，这些突变体材料可以用于玉米基因功能的研究和玉米育种。