Characterization of highly efficient heavy-ion mutagenesis in Arabidopsis thaliana.
Characterization of highly efficient heavy-ion mutagenesis in Arabidopsis thaliana.
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DOI：
10.1186/1471-2229-11-161
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发表时间：
2011-11-15
影响因子：
5.3
通讯作者：
Abe T
中科院分区：
文献类型：
--
作者：
Kazama Y;Hirano T;Saito H;Liu Y;Ohbu S;Hayashi Y;Abe T
重离子诱变是一种产生新突变体的有效技术，尤其是在高等植物中。重离子束具有高线性能量转移（LET），因此比其他诱变技术更有效地诱导DNA双链断裂。在此之前，我们通过分析LET对拟南芥突变诱导的影响，确定了最有效的重离子LET（LETmax：30.0 keV μm-1）。然而，LETmax的重离子诱导的突变DNA的分子结构仍不清楚。突变DNA结构的知识将有助于重离子束诱变的有效利用。用LETmax为400戈伊、LET为22.5 keV μm-1、剂量为250戈伊或450戈伊的碳（C）离子辐照拟南芥干种子。比较其对突变频率和DNA结构改变的影响。为了确定突变DNA的结构，我们筛选了下胚轴伸长（hy）和无毛（gl）的突变体，并通过高分辨率熔解曲线、PCR和测序分析鉴定了所得突变体中的突变DNA。LETmax时C离子诱发的突变频率是22.5 keV μm-1时的2倍，与甲磺酸乙酯诱发的突变频率相近。我们鉴定了22个突变DNA的结构。在两种LET中，超过80%的由C离子引起的突变是小于100 bp的碱基取代或缺失/插入。其他突变涉及大的重排。具有LETmax的C离子表现出高的突变效率，并且主要诱导碱基替换或小的缺失/插入，其中大多数是无效突变。这些小的改变可以通过单核苷酸多态性（SNP）检测系统来确定。因此，具有LETmax的C离子可用作与SNP检测系统结合的高效反向遗传系统，并且将有益于正向遗传学和植物育种。
Heavy-ion mutagenesis is recognised as a powerful technology to generate new mutants, especially in higher plants. Heavy-ion beams show high linear energy transfer (LET) and thus more effectively induce DNA double-strand breaks than other mutagenic techniques. Previously, we determined the most effective heavy-ion LET (LETmax: 30.0 keV μm-1) for Arabidopsis mutagenesis by analysing the effect of LET on mutation induction. However, the molecular structure of mutated DNA induced by heavy ions with LETmax remains unclear. Knowledge of the structure of mutated DNA will contribute to the effective exploitation of heavy-ion beam mutagenesis. Dry Arabidopsis thaliana seeds were irradiated with carbon (C) ions with LETmax at a dose of 400 Gy and with LET of 22.5 keV μm-1 at doses of 250 Gy or 450 Gy. The effects on mutation frequency and alteration of DNA structure were compared. To characterise the structure of mutated DNA, we screened the well-characterised mutants elongated hypocotyls (hy) and glabrous (gl) and identified mutated DNA among the resulting mutants by high-resolution melting curve, PCR and sequencing analyses. The mutation frequency induced by C ions with LETmax was two-fold higher than that with 22.5 keV μm-1 and similar to the mutation frequency previously induced by ethyl methane sulfonate. We identified the structure of 22 mutated DNAs. Over 80% of the mutations caused by C ions with both LETs were base substitutions or deletions/insertions of less than 100 bp. The other mutations involved large rearrangements. The C ions with LETmax showed high mutation efficiency and predominantly induced base substitutions or small deletions/insertions, most of which were null mutations. These small alterations can be determined by single-nucleotide polymorphism (SNP) detection systems. Therefore, C ions with LETmax might be useful as a highly efficient reverse genetic system in conjunction with SNP detection systems, and will be beneficial for forward genetics and plant breeding.
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影响因子：
1.1
作者：
Morita, Ryouhei;Kusaba, Makoto;Nishimura, Minoru
通讯作者：
Nishimura, Minoru
影响因子：
2.8
作者：
Masumura, K;Kuniya, K;Nohmi, T
通讯作者：
Nohmi, T
DOI：
10.1016/s0027-5107(98)00009-8
发表时间：
1998-06-05
影响因子：
2.3
作者：
Cecchini, E;Mulligan, BJ;Milner, JJ
通讯作者：
Milner, JJ
影响因子：
2.6
作者：
Höglund, E;Blomquist, E;Stenerlöw, B
通讯作者：
Stenerlöw, B
影响因子：
2.6
作者：
BLOCHER, D
通讯作者：
BLOCHER, D