重离子的辐射生物效应及其在生命科学中的应用

doi:10.13560/j.cnki.biotech.bull.1985.2017-0735

摘要/Abstract

摘要： 重离子是指重于元素周期表中2号元素氦并被电离的粒子。高能重离子因在其穿透物质的径迹上产生很强的局部电离,与传统的光子辐射（如X、γ射线）相比,会诱导更严重的辐射损伤生物效应。目前的机理研究显示,重离子辐射造成细胞DNA局部一到两个螺旋内出现多类型、多数量的损伤,即DNA团簇损伤。这种复杂的损伤影响DNA修复酶与DNA片段结合,进而影响修复,导致细胞死亡或产生不正确修复（即突变）。重离子辐射易引起突变的特征在植物与微生物诱变育种方面得到青睐。此外,重离子的细胞杀伤作用大,而且重离子具有倒转的剂量分布优势,将其用于癌症治疗,可使深部区域肿瘤细胞因高剂量辐射而被有效杀灭,而对周围组织因低剂量分布、相对危害较小,能实现精确靶向治疗,这被认为是最有前景的肿瘤放疗技术。对由重离子造成的DNA团簇损伤及其修复机制,以及两个重要修复途径的研究进展,进行了总结与深入分析。此外,我们还对重离子在辐射诱变育种和肿瘤治疗方面的应用进行了概要介绍,以期为从事重离子辐射生物学研究的人员理解DNA团簇损伤与修复机制及未来研究方向提供参考,促进对重离子辐射危害的评估与防护策略的建立、及其在生命科学中的良好应用。

关键词: 重离子, DNA团簇损伤, 辐射诱变育种, 重离子放射治疗

Abstract: The heavy ions are the ionized particles that are heavier than element He of second one in the element periodic table. Because of the strong local ionization on the pathway of penetration,the high-energy heavy ions induce the biological effects of more serious radiation damage compared with the traditional photon radiation,such as X-rays or γ-. The researches onmechanism show that radiation by heavy ions results in multiple-type and multiple-number damages at one or two helixes of DNA,i.e.,the clustered DNA damage. The complex damages impede the binding of DNA repair enzyme with DNA segment and the consequent repairing of cells,resulting in the subsequent death of cells or improper repair（i.e.,mutation）. The characteristic of heavy ion radiation,easily inducing mutant,is popular in mutation breeding of plant and microorganism. In addition,due to the high cell killing capability and the advantage of inverted dose profile,the exact targeted therapy can be achieved using heavy ions to treat tumor,because it delivers high doses of radiation to the deep regions of tumor cells to efficiently kill them but causes relatively little damages to the surround tissues at low dose;thus,heavy ion beam is deemed a most promising technology of tumor radiotherapy. Here we review and deeply analyze the clustered DNA damage induced by heavy ions and its repairing mechanism,as well as the research advances on the 2 key repair pathways. Besides,we summarize the application of heavy ions in mutation breeding and cancer therapy. It is expected that this review may provide a convenience for researchers in radiation biology of heavy ion to understand clustered DNA damage and repair mechanisms,and culture the research direction for better estimating the risks of heavy ions to human health and setting up better protection strategy as well as the application in life science.

Key words: heavy ions, clustered DNA damage, radiation-induced breeding, heavy ion radiotherapy

贾蓉, 苏锋涛, 胡步荣. 重离子的辐射生物效应及其在生命科学中的应用[J]. 生物技术通报, 2018, 34(1): 67-78.

JIA Rong, SU Feng-tao, HU Bu-rong. The Biological Effects Induced by Heavy Ion Radiation and Its Application in Life Science[J]. Biotechnology Bulletin, 2018, 34(1): 67-78.

参考文献

[1] 王同权, 沈永平, 王尚武, 等. 空间辐射环境中的辐射效应[J]. 国防科技大学学报, 1999, 21（4）：36-39.
[2] 葛本伟, 郭世先, 葛淼. 地球磁场与人体健康[J]. 国外医学（医学地理分册）, 2005, 26（1）：39-42.
[3] Blaisdell JO, Harrison L, Wallace SS. Base excision repair processing of radiation-induced clustered DNA lesions[J]. Radiation Protection Dosimetry, 2001, 97（1）：25-31.
[4] Goodhead DT, Thacker J, Cox R. Effects of radiations of different qualities on cells：molecular mechanisms of damage and repair[J]. Int J Radiat Biol, 1993, 63（5）：543-556.
[5] Dianov GL, O’Neill P, Goodhead DT. Securing genome stability by orchestrating DNA repair：removal of radiation-induced clustered lesions in DNA[J]. Bioessays, 2001, 23（8）：745-749.
[6] Brenner DJ, Ward JF. Constraints on energy deposition and target size of multiply damaged sites associated with DNA double-strand breaks[J]. Int J Radiat Biol, 1992, 61（6）：737-748.
[7] Okayasu R, Okada M, Okabe A, et al. Repair of DNA damage induced by accelerated heavy ions in mammalian cells proficient and deficient in the non-homologous end-joining pathway[J]. Radiation Research, 2006, 165（1）：59-67.
[8] Hamada N, Imaoka T, Masunaga S, et al. Recent advances in the biology of heavy-ion cancer therapy[J]. J Radiat Res, 2010, 51（4）：365-383.
[9] Heilmann J, Rink H, Taucher-scholz G, et al. DNA strand break induction and rejoining and cellular recovery in mammalian cells after heavy-ion irradiation[J]. Radiat Res, 1993, 135（1）：46-55.
[10] Tokuyama Y, Furusawa Y, Ideh, et al. Role of isolated and clustered DNA damage and the post-irradiating repair process in the effects of heavy ion beam irradiation[J]. J Radiat Res, 2015, 56（3）：446-455.
[11] Werner E, Wang Y, Doetsch PW. A single exposure to low-or high- LET radiation induces persistent genomic damage in mouse epithe-lial cells in vitro and in lung tissue[J]. Radiation Research, 2017. Doi： doi:10. 1667/RR14685. 1.
[12] Asaithamby A, Uematsu N, Chatterjee A, et al. Repair of HZE-particle-induced DNA double-strand breaks in normal human fibroblasts[J]. Radiat Res, 2008, 169（4）：437-446.
[13] Lorat Y, Timm S, Jakob B, et al. Clustered double-strand breaks in heterochromatin perturb DNA repair after high linear energy transfer irradiation[J]. Radiother Oncol, 2016, 121（1）：154-161.
[14] Taucher-scholz G, Heilmann J, Kraft G. Induction and rejoining of DNA double-strand breaks in CHO cells after heavy ion irradiation[J]. Adv Space Res, 1996, 18（1-2）：83-92.
[15] Tucker JD, Marples B, Ramsey MJ, et al. Persistence of chromosome aberrations in mice acutely exposed to 56 Fe +26 ions[J]. Radiation Research, 2004, 161（6）：648-655.
[16] Oike T, Niimi A, Okonogi N, et al. Visualization of complex DNA double-strand breaks in a tumor treated with carbon ion radiothe-rapy[J]. Scientific Reports, 2016, 6：22275. Doi： doi:10. 1038/ srep22275.
[17] Ritter S, Durante M. Durante M：Heavy-ion induced chromosomal aberrations：a review[J]. Mutation Research, 2010, 701（1）：38-46.
[18] Schipler A, Mladenova V, Soni A, et al. Chromosome thripsis by DNA double strand break clusters causes enhanced cell lethality, chromosomal translocations and 53BP1-recruitment[J]. Nucleic Acids Research, 2016, 44（16）：7673-7690.
[19] Asaithamby A, Hu B, Chen DJ. Unrepaired clustered DNA lesions induce chromosome breakage in human cells[J]. Proc Natl Acad Sci of the USA, 2011, 108（20）：8293-8298.
[20] Sutherland BM, Cuomo NC, Bennett PV. Induction of anchorage-independent growth in primary human cells exposed to protons or HZE ions separately or in dual exposures[J]. Radiation Research, 2005, 164（4）：493-496.
[21] Urushibara A. JAEA R&D Review, 2007, 71. （http://jolisfukyu. tokai-sc. jaea. go. jp/fukyu/mirai-en/2007/index_set. html）
[22] Shikazono N. JAEA R&D Review, 2008, 71（http://jolisfukyu. tokai-sc. jaea. go. jp/fukyu/mirai-en/2008/index_set. html）
[23] David-cordonnier MH, Cunniffe SM, Hickson ID, et al. Efficiency of incision of an AP site within clustered DNA damage by the major human AP endonuclease[J]. Biochemistry, 2002, 41（2）：634-642.
[24] Gulston M, Fulford J, Jenner T, et al. Clustered DNA damage induced by gamma radiation in human fibroblasts（HF19）, hamster（V79-4）cells and plasmid DNA is revealed as Fpg and Nth sensitive sites[J]. Nucleic Acids Research, 2002, 30（15）：3464-3472.
[25] Lomax ME, Salje H, Cunniffe S, et al. 8-OxoA inhibits the incision of an AP site by the DNA glycosylases Fpg, Nth and the AP endonuclease HAP1[J]. Radiation Research, 2005, 163（1）：79-84.
[26] Gulston M, Delara C, Jenner T, et al. Processing of clustered DNA damage generates additional double-strand breaks in mammalian cells post-irradiation[J]. Nucleic Acids Research, 2004, 32（4）：1602-1609.
[27] Blaisdell JO, Wallace SS. Abortive base-excision repair of radiation-induced clustered DNA lesions in Escherichia coli[J]. Proc Natl Acad Sci USA, 2001, 98（13）：7426-7430.
[28] D’Souza DI, Harrison L. Repair of clustered uracil DNA damages in Escherichia coli[J]. Nucleic Acids Research, 2003, 31（15）：4573-4581.
[29] Harrison L, Brame KL, Geltz LE, et al. Closely opposed apurinic/apyrimidinic sites are converted to double strand breaks in Escherichia coli even in the absence of exonuclease III, endonuclease IV, nucleotide excision repair and AP lyase cleavage[J]. DNA Repair, 2006, 5（3）：324-335.
[30] Malyarchuk S, Castore R, Harrison L. DNA repair of clustered lesions in mammalian cells：involvement of non-homologous end-joining[J]. Nucleic Acids Research, 2008, 36（15）：4872-4882.
[31] Fujimoto H, Pinak M, Nemoto T, et al. Molecular dynamics simulation of clustered DNA damage sites containing 8-oxoguanine and abasic site[J]. J Comput Chem, 2005, 26（8）：788-798.
[32] Sedelnikova OA, Redon CE, Dickey JS, et al. Role of oxidatively induced DNA lesions in human pathogenesis[J]. Mutation Research, 2010, 704（1-3）：152-159.
[33] Haines JW, Coster M, Bouffler SD. Impairment of the non-homologous end joining and homologous recombination pathways of DNA double strand break repair：Impact on spontaneous and radiation-induced mammary and intestinal tumour risk in Apc min/+ mice[J]. DNA Repair, 2015, 35：19-26.
[34] Jackson SP. Sensing and repairing DNA double-strand breaks [J]. Carcinogenesis, 2002, 23（5）：687-696.
[35] Wu X, Petrini JH, Heine WF, et al. Independence of R/M/N focus formation and the presence of intact BRCA1[J]. Science, 2000, 289（5476）：11.
[36] Cousineau I, Abaji C, Belmaaza A. BRCA1 regulates RAD51 function in response to DNA damage and suppresses spontaneous sister chromatid replication slippage：implications for sister chromatid cohesion, genome stability, and carcinogenesis[J]. Cancer Res, 2005, 65（24）：11384-11391.
[37] Fang S, Weissman AM. A field guide to ubiquitylation[J]. Cell Mol Life Sci, 2004, 61（13）：1546-1561.
[38] Jeggo PA. Identification of genes involved in repair of DNA double-strand breaks in mammalian cells[J]. Radiat Res, 1998, 150（5）：S80-91.
[39] Rothkamm K, Kruger I, Thompson LH, et al. Pathways of DNA double-strand break repair during the mammalian cell cycle[J]. Mol Cell Biol, 2003, 23（16）：5706-5715.
[40] Nagasawa H, Little JB, Inkret WC, et al. Response of X-ray-sensitive CHO mutant cells（xrs-6c）to radiation. II. Relationship between cell survival and the induction of chromosomal damage with low doses of alpha particles[J]. Radiat Res, 1991, 126（3）：280-288.
[41] Wang HY, Wang X, Zhang PY, et al. The Ku-dependent non-homologous end-joining but not other repair pathway is inhibited by high linear energy transfer ionizing radiation[J]. DNA Repair, 2008, 7（5）：725-733.
[42] Wang JH, Pluth JM, Cooper PK, et al. Artemis deficiency confers a DNA double-strand break repair defect and Artemis phosphorylation status is altered by DNA damage and cell cycle progression[J]. DNA Repair, 2005, 4（5）：556-570.
[43] Symington LS, Gautier J. Double-strand break end resection and repair pathway choice[J]. Annual Review of Genetics, 2011, 45：247-271.
[44] Couedel C, Mills KD, Barchi M, et al. Collaboration of homologous recombination and nonhomologous end-joining factors for the survival and integrity of mice and cells[J]. Genes Dev, 2004, 18（11）：1293-1304.
[45] Kass EM, Jasin M. Collaboration and competition between DNA double-strand break repair pathways[J]. Febs Letters, 2010, 584（17）：3703-3708.
[46] Shrivastav M, De haro LP, Nickoloff JA. Regulation of DNA double-strand break repair pathway choice[J]. Cell Res, 2008, 18（1）：134-147.
[47] You ZS, Bailis JM. CtIP coordinates DNA repair and cell cycle checkpoints[J]. Trends Cell Biol, 2010, 20（7）：402-409.
[48] Allen C, Kurimasa A, Brenneman MA, et al. DNA-dependent protein kinase suppresses double-strand break-induced and spontaneous homologous recombination[J]. Proc Natl Acad Sci USA, 2002, 99（6）：3758-3763.
[49] Pierce AJ, Hu P, Han MG, et al. Ku DNA end-binding protein modulates homologous repair of double-strand breaks in mammalian cells[J]. Genes Dev, 2001, 15（24）：3237-3242.
[50] Zafar F, Seidler SB, Kronenberg A, et al. Homologous recombina-tion contributes to the repair of DNA double-strand breaks induced by high-energy iron ions[J]. Radiat Res, 2010, 173（1）：27-39.
[51] Gerelchuluun A, Manabe E, Ishikawa T, et al. The major DNA repair pathway after both proton and carbon-Ion radiation is NHEJ, but the HR pathway is more relevant in carbon ions[J]. Radiat Res, 2015, 183（3）：345-356.
[52] Hammet A, Pike BL, Mcnees CJ, et al. FHA domains as phospho-threonine binding modules in cell signaling[J]. Iubmb Life, 2003, 55（1）：23-27.
[53] Durant ST, Nickoloff JA. Good timing in the cell cycle for precise DNA repair by BRCA1[J]. Cell Cycle, 2005, 4（9）：1216-1222.
[54] Deng CX. BRCA1：cell cycle checkpoint, genetic instability, DNA damage response and cancer evolution[J]. Nucleic Acids Res, 2006, 34（5）：1416-1426.
[55] Yan ZJ, Guo R, Paramasivam M, et al. A ubiquitin-binding protein, FAAP20, links RNF8-mediated ubiquitination to the Fanconi anemia DNA repair network[J]. Mol Cell, 2012, 47（1）：61-75.
[56] Cao L, Xu X, Bunting SF, et al. A selective requirement for 53BP1 in the biological response to genomic instability induced by Brca1 deficiency[J]. Mol Cell, 2009, 35（4）：534-541.
[57] Wang B, Matsuoka S, Carpenter PB, et al. 53BP1, a mediator of the DNA damage checkpoint[J]. Science, 2002, 298（5597）：1435-1438.
[58] Schultz LB, Chehab NH, MalikzaYA, et al. p53 binding protein 1（53BP1）is an early participant in the cellular response to DNA double-strand breaks[J]. J Cell Biol, 2000, 151（7）：1381-1390.
[59] 李仁民, 王菊芳, 李文建. 重离子束在微生物诱变育种及生物能源开发中的应用[J]. 原子核物理评论, 2007, 24（3）：234-237.
[60] Okamura M, Yasuno N, Ohtsuka M, et al. Wide variety of flower-color and -shape mutants regenerated from leaf cultures irradiatedwith ion beams[J]. Nucl Instrum Methods B, 2003, 206：574-578.
[61] Yamaguchi H, Nagatomi S, Morishita T, et al. Mutation induced with ion beam irradiation in rose[J]. Nucl Instrum Methods B, 2003, 206：561-564.
[62] Kikuchi S, Saito Y, Ryuto H, et al. Effects of heavy-ion beams on chromosomes of common wheat, Triticum aestivum[J]. Mutat Res, 2009, 669（1-2）：63-66.
[63] Li Q, Wei ZQ, Li WJ. Calculation of depth-dose distribution of intermediate energy heavy-ion beams[J]. Chinese Science Bulletin, 2002, 47（20）：1708-1710.
[64] 余增亮, 何建军, 邓建国, 等. 离子注入水稻诱变育种机理初探[J]. 安徽农业科学, 1989（1）：12-16.
[65] 郭高, 钱坤. 安徽省农科院育成世界上第一个离子束小麦新品种[J]. 安徽农业. l998, 25（5）：4.
[66] 卫增泉, 颉红梅, 梁剑平, 等. 重离子束在诱变育种和分子改造中的应用[J]. 原子核物理评论, 2003, 20（1）：38-41.
[67] 赵连芝, 壬勇, 甄东牛, 等. 春小麦突变新品种-“陇辐2号”[J]. 核农学报, 2005, 19（1）：80.
[68] 董喜存, 李文建, 何金玉, 等. 碳离子束辐照对甜高粱主要性状的影响[J]. 核技术, 2009, 32（2）：146-149.
[69] He JY. Pigment analysis of a color-leaf mutant in Wandering Jew（Tradescantia fluminensis）irradiated by carbon ions[J]. Nuclear Science and Techniques, 2011, 22（2）：77-83.
[70] Wu DL, Hou SW, Qian PP, et al. Flower color chimera and abnormal leaf mutants induced by 12 C 6+ heavy ions in Salvia splendens Ker-Gawl[J]. Sci Hortic, 2009, 121（4）：462-467.
[71] Yu LX, Li WJ, Du Y, et al. Flower color mutants induced by carbon ion beam irradiation of geranium（Pelargonium× hortorum, Bailey）[J]. Nuclear Science & Techniques, 2016, 27（5）：112-120.
[72] 周利斌, 李文建, 曲颖, 等. 重离子束辐照育种研究进展及发展趋势[J]. 原子核物理评论, 2008, 25（2）：165-170.
[73] Luo SW, Zhou B, Li WJ, et al. Mutagenic effects of carbon ion beam irradiations on dry Lotus japonicus seeds[J]. Nuclear Instruments & Methods in Physics Research Section B-beam Interactions with Materials and Atoms, 2016, 383（15）：123-128.
[74] Abe T, Matsuyana T, Shigeko S, et al. Chlorophyll-deficient mutants of rice demonstrated the deletion of a DNA fragment by heavy-ion irradiation[J]. J Radiat Res, 2002, 43（4）：S157-S161.
[75] Phanchaisri B, Samsang N, Yu LD, et al. Expression of OsSPY and 14-3-3 genes involved in plant height variations of ion-beam-induced KDML 105 rice mutants[J]. Mut Res, 2012, 734（1-2）：56-61.
[76] Tanaka A, Tano S, Chantes T, et al. A new Arabidopsis mutant induced by ion beams affects flavonoid synthesis with spotted pigmentation in testa[J]. Genes Genet Syst, 1997, 3（3）：141-148.
[77] Okamura M, Hase Y, Furusawa Y, et al. Tissue-dependent somaclonal mutation frequencies and spectra enhanced by ion beam irradiation in chrysanthemum[J]. Euphytica, 2015, 202（3）：333-343.
[78] Ishii K, Kazama Y, Morita R, et al. Linear Energy Transfer-Dependent Change in Rice Gene Expression Profile after Heavy-Ion Beam Irradiation[J]. PLoS One, 2016, 11（7）：e0160061.
[79] 都雯玥. 重离子辐照并筛选截短侧耳素高产菌株的研究[D]. 兰州：兰州理工大学, 2016：1-59.
[80] 颉红梅, 卫增泉, 李文建. 7 MeV/u O 6+ 离子对庆大霉素生产菌诱变的初步研究[J]. 辐射研究与辐射工艺学报, 1995, 13（2）：99-101.
[81] 王曙阳, 薄永恒, 王丽华, 等. 12 C 6+ 离子辐照对阿维链霉菌代谢效应研究[J]. 原子核物理评论, 2013, 30（2）：195-200.
[82] Li SW, Li M, Song HP, et al. Induction of a high-yield lovastatin mutant of Aspergillus terreus by 12 C 6+ heavy-ion beam irradiation and the influence of culture conditions on lovastatin production under submerged fermentation[J]. Applied Biochemistry and Biotechnology, 2011, 165（3）：913-925.
[83] Wang SY, Bo YH, Zhou X, et al. Significance of heavy-ion beam irradiation-induced avermectin b1a production by engineered streptomyces avermitilis[J]. Biomed Res Int, 2017, 2017：5373262. Doi： doi:10. 1155/2017/5373262.
[84] 胡伟, 陈积红, 张珍, 等. 重离子辐照柠檬酸菌株的诱变选育[J]. 辐射研究与辐射工艺学报, 2012, 30（1）：53-57.
[85] 严亚平, 王菊芳, 陆栋, 等. C离子束诱变产生甜高粱汁酒精酵母高产菌株的研究[J]. 原子核物理评论, 2009, 26（3）：269-273.
[86] Hu W, Chen JH, Li WJ, et al. Mutant breeding of Aspergillus niger irradiated by 12 C 6+ for hyper citric acid[J]. Nuclear Science and Techniques, 2014, 25（2）：1-4.
[87] 王雨辰, 王曙阳, 董妙音, 等. 重离子束辐照选育高产植物乳酸菌[J]. 辐射研究与辐射工艺学报, 2017, 35（1）：50-56.
[88] 李垄清, 马良, 李文建. 12 C 6+ 辐照环境下酵母高产β-葡聚糖菌株筛选及条件优化研究[J]. 食品科技, 2017, 42（3）：2-6.
[89] 薛林贵, 赵旭, 常思静, 等. 80 MeV/u C -12 离子诱变选育 PHB 高产菌株[J]. 核技术, 2010, 33（4）：284-288.
[90] Hu GR, Fan Y, Zhang L, et al. Enhanced lipid productivity and photosynthesis effi ciency in a Desmodesmus sp. mutant induced by heavy carbon ions[J]. PLoS One, 2013, 8（4）：e60700.
[91] 王芝瑶, 马玉彬, 牟润芝, 等. 重离子诱变创制高产油微拟球藻新品种[J]. 生物工程学报, 2013, 29（1）：119-122.
[92] Wang JF, Li RM, Lu D, et al. A quick isolation method for mutants with high lipid yield in oleaginous yeast[J]. World Journal of Microbiology and Biotechnology, 2009, 25（5）：921-925.
[93] Zhou X, Lu XH, Li XH, et al. Radiation induces acid tolerance of Clostridium tyrobutyricum and enhances bioproduction of butyric acid through a metabolic switch[J]. Biotechnology for Biofuels, 2014, 7：22.
[94] Suit H, Urie M. Proton beams in radiation therapy[J]. Journal of the National Cancer Institute, 1992, 84（3）：155-164.
[95] Linstadt DE, Castro JR, Phillips TL. Neon ion radiotherapy：results of the phase I/II clinical trial[J]. International Journal of Radiation Oncology Biology Physics, 1991, 20（4）：761-769.
[96] Castro JR. Results of Heavy-Ion Radiotherapy[J]. Radiat Environ Bioph, 1995, 34（1）：45-48.
[97] Combs SE, Jakel O, Haberer T, et al. Particle therapy at the heidelberg ion therapy center（HIT）- integrated research-driven university-hospital-based radiation oncology service in Heidelberg, Germany[J]. Radiother Oncol, 2010, 95（1）：41-44.
[98] Sawada K, Sawada J, Sakata T, et al. Performance test of electron cyclotron resonance ion sources for the Hyogo Ion Beam Medical Center[J]. Rev Sci Instrum, 2000, 71（2）：987-989.
[99] 辻进博彦. 日本重离子治疗临床研究[C]. 兰州：第二届全国重离子治疗高端论坛, 2016. 12. 27.
[100] 王小虎. 国产碳离子治癌设备应用进展[C]. 兰州：第二届全国重离子治疗高端论坛, 2016. 12. 27.
[101] Tsujii H, Kamada T. A review of update clinical results of carbon ion radiotherapy[J]. Jpn J Clin Oncol, 2012, 42（8）：670-685.
[102] Kitagawa A, Fujita T, Muramatsu M, et al. Review on heavy ion radiotherapy facilities and related ion sources（invited）[J]. Rev Sci Instrum, 2010, 81（2）：02B909.
[103] Sunada S, Cartwright IM, Hirakawa H, et al. Investigation of the relative biological effectiveness and uniform isobiological killing effects of irradiation with a clinical carbon SOBP beam on DNA repair deficient CHO cells[J]. Oncol Lett, 2017, 13（6）：4911-4916.
[104] Hada M, Sutherland BM. Spectrum of complex DNA damages depends on the incident radiation[J]. Radiation Research, 2006, 165（2）：223-230.
[105] Sunada S, Hirakawa H, Fujimori A, et al. Oxygen Enhancement ratio in radiation-induced initial DSBs by an optimized flow cytometry-based gamma-H2AX analysis in A549 human cancer cells[J]. Radiat Res, 2017, 188（5）：591-594.
[106] Cui X, Oonishi K, Tsujii H, et al. Effects of carbon ion beam on putative colon cancer stem cells and its comparison with X-rays[J]. Cancer Research, 2011, 71（10）：3676-3687.
[107] Osama M, Brock JS, Janapriya S, et al. Carbon ion radiotherapy：a review of clinical experiences and preclinical research, with an emphasis on DNA damage/repair[J]. Cancers（Basel）, 2017, 9（6）. pii：E66. Doi： doi:10. 3390/cancers9060066.
[108] Kuscu C, Parlak M, Tufan T, et al. CRISPR-STOP：gene silencing through base-editing-induced nonsense mutations[J]. Nat Methods, 2017, 14（7）：710-712.
[109] Sattar MN, Iqbal Z, Tahir MN, et al. CRISPR/Cas9：A practical approach in date palm genome editing[J]. Front Plant Sci, 2017, 8：1469.
[110] Huang JJ, Wang YF, Zhao JG. CRISPR editing in biological and biomedical investigation[J]. J Cell Physiol, 2017. doi:10. 1002/jcp. 26141.
[111] Donohoue PD, Barrangou R, May AP. Advances in industrial biotechnology using CRISPR-Cas systems[J]. Trends Biotechnol, 2017, pii：S0167-7799（17）30187-7.