Mature seed-derived callus of the model indica rice variety Kasalath is highly competent in Agrobacterium-mediated transformation.
Mature seed-derived callus of the model indica rice variety Kasalath is highly competent in Agrobacterium-mediated transformation.
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DOI：
10.1007/s00299-010-0921-x
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发表时间：
2010-12
影响因子：
6.2
通讯作者：
Toki, Seiichi
中科院分区：
文献类型：
--
作者：
Saika, Hiroaki;Toki, Seiichi
我们以前建立了一种高效的农杆菌介导的转化系统，该系统利用日本晴成熟种子的原代愈伤组织进行转化。我们预计，缩短的组织培养周期将减少愈伤组织褐变--在未分化状态下延长组织培养期间，这是籼型转化系统的常见问题。在本研究中，我们成功地将我们的高效转化体系应用于水稻模式品种Kasalath。LUC报告系统具有足够的灵敏度，可以定量分析水稻愈伤组织对农杆菌介导的转化能力。我们意外地发现，与日本晴相比，Kasalath的原代愈伤组织表现出非常高的农杆菌介导性转化能力。Southern杂交和Luc荧光分析表明，Kasalath的原代愈伤组织中成功地发生了独立的转化事件，其频率是日本晴的10倍，其中约40%的事件发生了单拷贝T-DNA整合。我们还比较了日本晴和Kasalath的次生愈伤组织的能力，再次发现Kasalath的次生愈伤组织的能力更强，尽管由于转化细胞和未转化细胞的旺盛生长，次级愈伤组织中独立的转化事件的鉴定和后续观察是困难的。Kasalath的高效转化系统有助于QTL基因的鉴定，因为许多QTL基因是在日本晴×Kasalath的遗传背景下分析的。Kasalath较高的转化能力可能是建立涉及基因打靶等新转化技术的高效系统的有用特征。本文的在线版本(doi：10.1007/s00299010921-x)包含补充材料，授权用户可以使用。
We previously established an efficient Agrobacterium-mediated transformation system using primary calli derived from mature seeds of the model japonica rice variety Nipponbare. We expected that the shortened tissue culture period would reduce callus browning—a common problem with the indica transformation system during prolonged tissue culture in the undifferentiated state. In this study, we successfully applied our efficient transformation system to Kasalath—a model variety of indica rice. The Luc reporter system is sensitive enough to allow quantitative analysis of the competency of rice callus for Agrobacterium-mediated transformation. We unexpectedly discovered that primary callus of Kasalath exhibits a remarkably high competency for Agrobacterium-mediated transformation compared to Nipponbare. Southern blot analysis and Luc luminescence showed that independent transformation events in primary callus of Kasalath occurred successfully at ca. tenfold higher frequency than in Nipponbare, and single copy T-DNA integration was observed in ~40% of these events. We also compared the competency of secondary callus of Nipponbare and Kasalath and again found superior competency in Kasalath, although the identification and subsequent observation of independent transformation events in secondary callus is difficult due to the vigorous growth of both transformed and non-transformed cells. An efficient transformation system in Kasalath could facilitate the identification of QTL genes, since many QTL genes are analyzed in a Nipponbare × Kasalath genetic background. The higher transformation competency of Kasalath could be a useful trait in the establishment of highly efficient systems involving new transformation technologies such as gene targeting. The online version of this article (doi:10.1007/s00299-010-0921-x) contains supplementary material, which is available to authorized users.
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影响因子：
2.4
作者：
Katagiri, S;Wu, JZ;Sasaki, T
通讯作者：
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影响因子：
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作者：
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