An Efficient System for Ds Transposon Tagging in Brachypodiurn distachyon
An Efficient System for Ds Transposon Tagging in Brachypodiurn distachyon
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二穗短柄藻 Ds 转座子标记的高效系统
DOI：
10.1104/pp.18.00875
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发表时间：
2019
期刊：
影响因子：
7.4
通讯作者：
An Hailong
中科院分区：
文献类型：
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作者：
Wu Hongyu;Xue Xiaodong;Qin Caihua;Xu Yi;Guo Yuyu;Li Xiang;Lv Wei;Li Qinxia;Mao Chuangxue;Li Luzhao;Zhao Suzhen;Qi Xiaoquan;An Hailong
转座子标记是一种强大的工具，已广泛应用于多种植物的插入诱变。一些研究的目标是在单子叶植物brachypodium disachyon中创建转移dna （T-DNA）插入突变群体，该植物被用作温带谷物研究的模型系统，但缺乏针对转座子策略的研究。在这里，我们描述了玉米（Zea mays）解离（Ds）转座子标记系统在b .distachyon中的应用。在相同的T-DNA中构建35s:: actpascassette和dselement，并将其转化为ob .distachyonplant。在激活因子（Ac）转座酶的作用下，该元素很容易转座到其他染色体或同一染色体上。通过同源染色体突触、重组和分离，元件从元件中分离出来。我们使用G418和GFP技术选择了只稳定的植株，并分析了241个株系，其中一些株系在产生只稳定的后代方面效率很高。通过热不对称交错PCR，我们从单基因型植物中分离出710个独立的侧翼序列。此外，我们通过这种转座子标记系统鉴定了大量具有可见发育表型的突变体。与传统的T-DNA插入策略相比，该系统相对简单和快速，因为一旦获得高效系，它们可以重复使用，从非转座植物中产生更多的系，而无需使用耗时的组织培养步骤。
Transposon tagging is a powerful tool that has been widely applied in several species for insertional mutagenesis in plants. Several efforts have aimed to create transfer-DNA (T-DNA) insertional mutant populations inBrachypodium distachyon, a monocot plant used as a model system to study temperate cereals, but there has been a lack of research aimed at using transposon strategies. Here, we describe the application of a maize (Zea mays)Dissociation(Ds) transposon tagging system inB.distachyon. The35S::AcTPasecassette andDselement were constructed within the same T-DNA and transformed intoB.distachyonplants. TheDselement was readily transposed to other chromosomes or to the same chromosome under the function ofActivator(Ac) transposase. Through homologous chromosome synapsis, recombination, and segregation, theDselement separated from theAcelement. We selected stableDs-only plants using G418 and GFP assays and analyzed 241 T0lines, some of which were highly efficient at producingDs-only progeny. Through thermal asymmetric interlaced PCR, we isolated 710 independentDsflanking sequences fromDs-only plants. Furthermore, we identified a large collection of mutants with visible developmental phenotypes via this transposon tagging system. The system is relatively simple and rapid in comparison to traditional T-DNA insertion strategies, because once efficiency lines are obtained they can be reused to generate more lines from nontransposed plants without the use of time-consuming tissue culture steps.