Effect of high-LET Fe-ion beam irradiation on mutation induction in &lt;i&gt;Arabidopsis thaliana&lt;/i&gt;

Yusuke Kazama,Yuki Shirakawa,Tomoko Abe,Tomonari Hirano,Kiyoshi Nishihara,Sumie Ohbu

doi:10.1266/ggs.88.189

Abstract

Heavy-ion beams are powerful mutagens. They cause a broad spectrum of mutation phenotypes with high efficiency even at low irradiation doses and short irradiation times. These mutagenic effects are due to dense ionisation in a localised region along the ion particle path. Linear energy transfer (LET; keV·μm(-1)), which represents the degree of locally deposited energy, is an important parameter in heavy-ion mutagenesis. For high LET radiation above 290 keV∙μm(-1), however, neither the mutation frequency nor the molecular nature of the mutations has been fully characterised. In this study, we investigated the effect of Fe-ion beams with an LET of 640 keV∙μm(-1) on both the mutation frequency and the molecular nature of the mutations. Screening of well-characterised mutants (hy and gl) revealed that the mutation frequency was lower than any other ion species with low LET. We investigated the resulting mutations in the 4 identified mutants. Three mutants were examined by employing PCR-based methods, one of which had 2-bp deletion, another had 178 bp of tandemly duplication, and other one had complicated chromosomal rearrangements with variable deletions in size at breakpoints. We also detected large deletions in the other mutant by using array comparative genomic hybridisation. From the results of the analysis of the breakpoints and junctions of the detected deletions, it was revealed that the mutants harboured chromosomal rearrangements in their genomes. These results indicate that Fe-ion irradiation tends to cause complex mutations with low efficiency. We conclude that Fe-ion irradiation could be useful for inducing chromosomal rearrangements or large deletions.

Highlights

Heavy-ion beams comprise accelerated heavy ions that are produced by using accelerators such as cyclotrons or synchrotrons
Mutant screening The hy and gl mutants were screened in the M2 generation after Fe-ion irradiation at a dose of 50 Gy, for which the highest frequencies of albino mutants was observed in the dose range 5–150 Gy (Kazama et al, 2008)
Characterisation of DNA mutations induced by using Fe-ion irradiation To investigate the structure of mutated DNA in the isolated mutants that was caused by carrying out Fe-ion irradiation, DNA from the isolated mutants was subjected to PCR and sequencing analyses by using primers specific for the genes that were responsible for the hy, gl, bp, and nl phenotypes

Summary

Introduction

Heavy-ion beams comprise accelerated heavy ions that are produced by using accelerators such as cyclotrons or synchrotrons. They have been accepted as an efficient technology for the mutation breeding of plants and microbes. Studies on biological processes in plants and microbes have been advanced by using heavy-ion irradiation (Hase et al, 2000, 2006; Shitsukawa et al, 2007; Shimada et al, 2009; Yasui et al, 2011; Fujita et al, 2012; Sasaki et al, 2012; Ishikawa et al, 2012; Kazama et al, 2013; Hirano et al, 2013; Ma et al, 2013). Heavy-ion beams are called high linear energy transfer (LET; keV·μm–1) irradiation, as opposed to low LET irradiation from X-rays and γ-rays. When the albino mutant incidence was investigated in the M2 generation after irradiation treatment with heavy-ion beams with LETs ranging from 22.5 to 640

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Discussion

Conclusion