Bhatt A M, Page T, Lawson E J, Lister C, Dean C
Department of Molecular Genetics, John Innes Centre, Norwich, UK.
Plant J. 1996 Jun;9(6):935-45. doi: 10.1046/j.1365-313x.1996.9060935.x.
A pilot-scale transposon mutagenesis experiment using a modified autonomous Activator (Ac) element, Ac delta Nael, was carried out in Arabidopsis thaliana. Four different transformants carrying Ac elements in different and defined genomic locations were used to generate 1000 plants carrying approximately 500 independent germinal transposition events. These plants were then selfed and the 1000 families screened in tissue culture and soil for phenotypic mutants. Fifty different families segregated mutations in their progeny. Preliminary Southern blot analysis of 29 families which segregated mutant progeny, showed that 28 had a transposed Ac. Six of the families were further tested for linkage between the transposed Ac and the mutant phenotype, and instability of the putatively tagged locus. Two of the mutants were shown to be tagged as they were tightly linked to a transposed Ac, and somatic and germinal reversion was associated with loss of Ac. One other mutant locus was shown to be closely linked to a transposed Ac, and therefore was likely to be tagged. The remaining three mutations were not tagged as they were not linked to a transposed Ac. In two of the tagged mutants Ac had transposed to closely linked sites, while in a third mutant the co-segregating Ac had transposed to a site which was not tightly linked to the donor T-DNA. Multiple insertions into the DIF1 locus were found, due to the preferential transposition of Ac to a linked site.
利用一种经过修饰的自主激活子（Ac）元件Ac delta Nael，在拟南芥中开展了一项中试规模的转座子诱变实验。使用了四个在不同且明确的基因组位置携带Ac元件的不同转化体，来培育1000株携带约500个独立生殖转座事件的植株。然后让这些植株自交，并在组织培养和土壤中筛选这1000个家系中的表型突变体。五十个不同的家系在其后代中分离出了突变。对29个分离出突变后代的家系进行的初步Southern杂交分析表明，其中28个家系有一个转座的Ac。对其中六个家系进一步检测了转座的Ac与突变表型之间的连锁关系，以及假定被标记位点的不稳定性。其中两个突变体被证明是被标记的，因为它们与一个转座的Ac紧密连锁，并且体细胞和生殖细胞回复与Ac的丢失相关。另一个突变位点被证明与一个转座的Ac紧密连锁，因此很可能被标记。其余三个突变没有被标记，因为它们与一个转座的Ac没有连锁关系。在两个被标记的突变体中，Ac转座到了紧密连锁的位点，而在第三个突变体中，共分离的Ac转座到了一个与供体T-DNA不紧密连锁的位点。由于Ac优先转座到一个连锁位点，发现了多个插入到DIF1位点的情况。
