Brachypodium distachyon is an annual C3 grass used as a monocot model system in functional genomics research. Insertional mutagenesis is a powerful tool for both forward and reverse genetics studies. In this study, we explored the possibility of using the tobacco retrotransposon Tnt1 to create a transposon‐based insertion mutant population in B. distachyon. We developed transgenic B. distachyon plants expressing Tnt1 (R0) and in the subsequent regenerants (R1) we observed that Tnt1 actively transposed during somatic embryogenesis, generating an average of 6.37 insertions per line in a population of 19 independent R1 regenerant plants analyzed. In seed‐derived progeny of R1 plants, Tnt1 segregated in a Mendelian ratio of 3:1 and no new Tnt1 transposition was observed. A total of 126 flanking sequence tags (FSTs) were recovered from the analyzed R0 and R1 lines. Analysis of the FSTs showed a uniform pattern of insertion in all the chromosomes (1–5) without any preference for a particular chromosome region. Considering the average length of a gene transcript to be 3.37 kb, we estimated that 29 613 lines are required to achieve a 90% possibility of tagging a given gene in the B. distachyon genome using the Tnt1‐based mutagenesis approach. Our results show the possibility of using Tnt1 to achieve near‐saturation mutagenesis in B. distachyon, which will aid in functional genomics studies of other C3 grasses.

中文翻译：

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使用Tnt1可逆转座子元素诱变短枝芽孢杆菌的诱变。

Brachypodium distachyon是一年生的C3草，在功能基因组学研究中用作单子叶植物模型系统。插入诱变是进行正向和反向遗传学研究的有力工具。在这项研究中，我们探讨了使用烟草反转录转座子的可能性TNT1建立在基于转座子插入突变体人口二穗短柄草。我们开发了表达Tnt1（R0）的转基因B. distachyon植物，在随后的再生子（R1）中，我们观察到Tnt1在体细胞胚发生过程中被主动转座，在分析的19个独立R1再生植物群体中平均每株产生6.37个插入。在R1植物的种子后代中，Tnt1以3：1的孟德尔比率分离，未观察到新的Tnt1转座。从分析的R0和R1品系中总共回收了126个侧翼序列标签（FST）。对FST的分析显示，所有染色体（1-5）的插入方式均一，而对特定染色体区域没有任何偏好。考虑一个基因转录物的平均长度为3.37 kb的，我们估计，29条613线都需要实现在标记的给定基因的90％可能性二穗短柄草使用基因组TNT1基础的诱变方法。我们的结果表明，使用Tnt1可以实现B. distachyon的近饱和诱变，这将有助于其他C3草的功能基因组学研究。