Efficient methods for multigene transformation are important for developing novel crop varieties. Methods based on random integrations of multiple genes have been successfully used for metabolic engineering in plants. However, efficiency of co‐integration and co‐expression of the genes could present a bottleneck. Recombinase‐mediated integration into the engineered target sites is arguably a more efficient method of targeted integration that leads to the generation of stable transgenic lines at a high rate. This method has the potential to streamline multigene transformation for metabolic engineering and trait stacking in plants. Therefore, empirical testing of transgene(s) stability from the multigene site‐specific integration locus is needed. Here, the recombinase technology based on Cre‐lox recombination was evaluated for developing multigenic lines harboring constitutively‐expressed and inducible genes. Targeted integration of a five genes cassette in the rice genome generated a precise full‐length integration of the cassette at a high rate, and the resulting multigenic lines expressed each gene reliably as defined by their promoter activity. The stable constitutive or inducible expression was faithfully transmitted to the progeny, indicating inheritance‐stability of the multigene locus. Co‐localization of two distinctly inducible genes by heat or cold with the strongly constitutive genes did not appear to interfere with each other's expression pattern. In summary, high rate of co‐integration and co‐expression of the multigene cassette installed by the recombinase technology in rice shows that this approach is appropriate for multigene transformation and introduction of co‐segregating traits.

中文翻译：

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水稻中重组酶介导的多基因盒整合导致堆叠基因座的稳定表达和遗传。

多基因转化的有效方法对于开发新的作物品种很重要。基于多个基因随机整合的方法已成功用于植物代谢工程。然而，基因的共整合和共表达的效率可能会成为瓶颈。重组酶介导的整合到工程靶位点可以说是一种更有效的靶向整合方法，可导致以高速率产生稳定的转基因系。这种方法有可能简化植物代谢工程和性状堆叠的多基因转化。因此，需要对来自多基因位点特异性整合基因座的转基因稳定性进行实证测试。在这里，基于 Crelox 的重组酶技术评估了重组以开发含有组成型表达和诱导基因的多基因系。水稻基因组中五个基因盒的靶向整合以高速率产生了该盒的精确全长整合，并且由此产生的多基因系可靠地表达了由其启动子活性定义的每个基因。稳定的组成型或诱导型表达忠实地传递给后代，表明多基因位点的遗传稳定性。两个通过热或冷明显诱导的基因与强组成基因的共定位似乎不会干扰彼此的表达模式。总之，