Rapid, large-scale generation of Ds transposant lines and analysis of the Ds insertion sites in rice
Rapid, large-scale generation of Ds transposant lines and analysis of the Ds insertion sites in rice
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DOI：
10.1111/j.1365-313x.2004.02116.x
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发表时间：
2004-07-01
期刊：
影响因子：
7.2
通讯作者：
Han, CD
中科院分区：
文献类型：
--
作者：
Kim, CM;Piao, HL;Han, CD
快速，大规模的一代的Ds转座群体实现了使用再生程序，涉及组织培养的种子来源的愈伤组织携带Ac和非活性Ds元素。在来自相同Ds和Ac起始系之间的遗传杂交的F-2后代中，大多数杂交产生了10- 20%的独立的germinal转座频率。此外，许多Ds元件进行固定化，即使Ac的表达。通过比较，在愈伤组织衍生的再生群体中，超过70%的植物进行独立的Ds插入，表明转座早期愈伤组织形成。在剩余的种群中，大多数植物仅携带Ac。大多数新的Ds插入稳定地传递到下一代。再生群体中独立转座子的比例异常高意味着可能不一定需要转座Ds的选择标记和Ac/Ds活性的持续监测。通过对1297个Ds侧翼序列的分析，构建了一张包含1072个Ds插入位点的遗传图谱。图谱显示Ds元件被转座到所有的水稻染色体上，不仅在供体位点附近（36%），而且在某些物理上不连锁的臂上也有偏好。遗传杂交和组织培养的群体显示了相同的Ds插入位点的分布模式。这些Ds插入位点的信息已存入GenBank。其中，55%的Ds元件位于预测的开放阅读框（ORF）区域。因此，我们提出了一个最佳的策略，在水稻中快速产生大量的Ds转座子。
Rapid, large-scale generation of a Ds transposant population was achieved using a regeneration procedure involving tissue culture of seed-derived calli carrying Ac and inactive Ds elements. In the F-2 progeny from genetic crosses between the same Ds and Ac starter lines, most of the crosses produced an independent germinal transposition frequency of 10-20%. Also, many Ds elements underwent immobilization even though Ac was expressed. By comparison, in a callus-derived regenerated population, over 70% of plants carried independent Ds insertions, indicating transposition early in callus formation. In the remaining population, the majority of plants carried only Ac. Most of the new Ds insertions were stably transmitted to a subsequent generation. An exceptionally high proportion of independent transposants in the regenerated population means that selection markers for transposed Ds and continual monitoring of Ac/Ds activities may not necessarily be required. By analyzing 1297 Ds-flanking DNA sequences, a genetic map of 1072 Ds insertion sites was developed. The map showed that Ds elements were transposed onto all of the rice chromosomes, with preference not only near donor sites (36%) but also on certain physically unlinked arms. Populations from both genetic crossing and tissue culture showed the same distribution patterns of Ds insertion sites. The information of these mapped Ds insertion sites was deposited in GenBank. Among them, 55% of Ds elements were on predicted open-reading frame (ORF) regions. Thus, we propose an optimal strategy for the rapid generation of a large population of Ds transposants in rice.