JoVE Journal > Immunology and Infection
Summary
Abstract
Introduction
Protocol
Results
Discussion
Disclosures
Acknowledgements
Materials
References
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Immunology and Infection

ラムダを選択cII変異検出システム

Published: April 26, 2018
doi:
10.3791/57510
,
1Department of Preventive Medicine, USC Keck School of Medicine,University of Southern California

Summary

トランスジェニックげっ歯類の培養細胞でラムダを選択cII突然変異試験または興味の化学/物理エージェントで治療対応する動物のための詳しいプロトコルについて述べる。このアプローチは、哺乳類細胞における発がん性物質の変異原性試験のため広く使用されています。

Abstract

トランスジェニック動物モデルおよび突然変異検出システムの数は、哺乳類細胞における発がん性物質の変異原性試験のために開発されています。これらのトランスジェニック マウスとラムダ (λ) の選択cII変異検出システム世界中の多くの研究グループによって変異原性実験のため採用されています。ここでは、ラムダを選択cII突然変異試験は、トランスジェニック マウス/ラットの培養細胞に適用することができるための詳しいプロトコルについて述べるまたは興味の化学/物理エージェントに対応する動物が扱われます。プロトコルは、次の手順で構成されます: (1) 分離細胞からゲノム dna やトランスジェニック動物の臓器・組織生体外でまたは生体内で、それぞれで治療テスト混合物;(ゲノム DNA からの (すなわち、 cII遺伝子) の変異レポーター遺伝子を運ぶラムダ シャトルベクターの回復 2)(伝染性のバクテリオファージに救助されたベクトルの包装 3)(4) ホストの細菌に感染して誘導cII突然変異の伝播を許可する選択的な条件下で培養(5) 得点cII-変異体と DNA シーケンスをそれぞれcII突然変異頻度および突然変異スペクトルを分析します。

Introduction

トランスジェニック動物モデルおよび突然変異検出システムの広い範囲は、哺乳類細胞における発がん性物質の変異原性試験のために開発されています。これらの大きなブルー (以下 bb) トランスジェニック マウスと λ 選択cII変異検出システムで採用されている変異原性実験のためこのグループは、多くの他研究グループ世界中1,の2、 3,4,5,6,7,8,9。過去 16 年間、これらのトランスジェニック動物を用いた化学的および/または物理的な剤の変異原性の効果を調べたか、対応する胚性線維芽細胞培養テスト混合物と扱われ、その後の分析、λ 選択cIIアッセイおよび DNA の配列、それぞれ10、11,12,の13,14によってcII遺伝子の遺伝子型と表現型,15,16,17,18,19,20,21,22,23,24. これらのトランスジェニック動物のゲノムに複数コピー頭-尾コンカテマー1,2,25, 染色体 4 統合バクテリオファージ λ シャトル ベクトル (λLIZ) が含まれています。ΛLIZ シャトルベクターを運ぶ 2 つの変異レポーター遺伝子、すなわち、ラッチとcII遺伝子1,2,25,26,27、 28,29,30,31,32,33,34,35,36、 37,38,39,40,41,42,43,44,45,46,47. λ 選択cIIアッセイに基づいて λLIZ シャトル ベクトルのトランスジェニック動物1,2,25 の臓器・組織由来細胞のゲノム DNA からの回復.回復 λLIZ シャトル ベクトル、インジケーター ホスト大腸菌に感染可能な λ ファージの頭にパッケージ化されます。その後、感染した細菌は、得点とcII遺伝子1,3における変異解析を可能にする選択的な条件のもとで栽培しています。

ここ、体外治療トランスジェニック動物の細胞・臓器からゲノム DNA の隔離から成っている λ 選択cIIの試金のための詳しいプロトコルについて述べる/シャトル生体内でλLIZ の化合物は、取得テスト ベクトルgenomic DNA から、バクテリオファージ、 cIIの同定とホストエシェリヒア属大腸菌の感染感染 λ ファージにベクトルの包装- cIIの突然変異頻度を決定する選択的な条件の下で突然変異体とCII変異スペクトルを確立する dna 塩基配列解析。トランスジェニック マウス ・ ラット細胞培養体外興味の化学/物理エージェントに適用できるプロトコルまたは対応する動物の組織/臓器治療体内テスト化学/エージェント1、 2,4,48,49,50,51,52。Λ 選択cIIアッセイの図式を図 1に示します。

Protocol

1 マウス胚性線維芽細胞からゲノム DNA の隔離 注: プライマリ マウス胚性線維芽細胞は、公開されたプロトコル53によると BB C57BL/6 遺伝的背景を持つトランスジェニック マウス由来の胚から分離されます。このプロトコルのための開始材料は、1 x 106 1 x 107胚性線維芽細胞細胞制御化合物とテストで構成されます。収穫と標準的な方?…

Representative Results

データ ディストリビューションによってパラメトリックまたはノンパラのテストを使用して、処置および制御グループ (すなわち、自発の突然変異体の周波数対誘導) のcIIの突然変異頻度の違いの重要性を判断します.誘導cII変異周波数別の治療グループ間での比較が行われます各種 (一対)、適用可能な統計的テスト。Χ2テストや分散分析 (…

Discussion

Λ 選択cIIアッセイは、3BB の齧歯動物の臓器・組織由来細胞のゲノム DNA から回復したcII遺伝子変異の検出に使用されます。これらの形質転換動物ゲノムには染色体統合 λLIZ シャトル ベクトルは、 cIIを運ぶ複数タンデム コピーが含まれます (294 bp) とラッチ(1,080 bp) 遺伝子変異レポーター遺伝子1,2,</su…

Disclosures

The authors have nothing to disclose.

Acknowledgements

すべての同僚の貢献と (説明のため) この原稿に呼ばれている、結果が私たちの元の研究に協力したいと思います。作家の作品は、国立歯科・頭蓋顔面研究、国立衛生研究所 (1R01DE026043) ab の AB (California Tobacco-Related 病研究プログラムの大学からの補助金によってサポートされてTRDRP-26IR-0015) および ST (TRDRP-25IP-0001)。研究のスポンサーには、研究デザイン、データ収集、データ分析、データ解釈、レポートの執筆、出版のために提出する決定の役割はなかった。

Materials

Agar MO Bio Laboratories, Inc. 12112-05 Bacteriological grade
BigDye Terminator v3.1 Cycle Sequencing Kit Thermo Fisher Scientific 4337455 None
Casein Peptone Alfa Aesar H26557 None
Gelatine J. T. Baker 2124-01 Powder
Glycerol Fisher Scientific BP 229-1 / M-13750 None
LB Agar Fisher Scientific BP 9724-500 None
QIAquick PCR purification kit Qiagen 8104 50 PCR purification reactions
Sodium Acetate Trihydrate Fisher Scientific M-15756 None
Taq5000 DNA Polymerase  Qiagen 201207 None
Thiamine Hydrochloride Macron Fine Chemicals 2722-57 None
Transpack Packaging Extract Stratagene Corp., Acquired by BioReliance | Sigma-Aldrich Corp. 200223 50 packaging reactions
Tris Base Fisher Scientific BP 152-1 / EC 201-064-4 None
Trypton Biosciences RC-110 None
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References

  1. Jakubczak, J. L., et al. Analysis of genetic instability during mammary tumor progression using a novel selection-based assay for in vivo mutations in a bacteriophage lambda transgene target. Proc Natl Acad Sci U S A. 93 (17), 9073-9078 (1996).
  2. Lambert, I. B., Singer, T. M., Boucher, S. E., Douglas, G. R. Detailed review of transgenic rodent mutation assays. Mutat Res. 590 (1-3), 1-280 (2005).
  3. Besaratinia, A., Pfeifer, G. P. Investigating human cancer etiology by DNA lesion footprinting and mutagenicity analysis. Carcinogenesis. 27 (8), 1526-1537 (2006).
  4. Watson, D. E., Cunningham, M. L., Tindall, K. R. Spontaneous and ENU-induced mutation spectra at the cII locus in Big Blue Rat2 embryonic fibroblasts. Mutagenesis. 13 (5), 487-497 (1998).
  5. Erexson, G. L., Watson, D. E., Tindall, K. R. Characterization of new transgenic Big Blue(R) mouse and rat primary fibroblast cell strains for use in molecular toxicology studies. Environ Mol Mutagen. 34 (2-3), 90-96 (1999).
  6. Erexson, G. L., Tindall, K. R. Micronuclei and gene mutations in transgenic big Blue((R)) mouse and rat fibroblasts after exposure to the epoxide metabolites of 1, 3-butadiene. Mutat Res. 472 (1-2), 105-117 (2000).
  7. McDiarmid, H. M., Douglas, G. R., Coomber, B. L., Josephy, P. D. 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced mutagenesis in cultured Big Blue rat mammary epithelial and fibroblast cells. Environ Mol Mutagen. 39 (2-3), 245-253 (2002).
  8. Papp-Szabo, E., Douglas, G. R., Coomber, B. L., Josephy, P. D. Mutagenicity of the oral carcinogen 4-nitroquinoline-1-oxide in cultured BigBlue rat tongue epithelial cells and fibroblasts. Mutat Res. 522 (1-2), 107-117 (2003).
  9. Guichard, Y., et al. In Vitro Study of Mutagenesis Induced by Crocidolite-Exposed Alveolar Macrophages NR8383 in Cocultured Big Blue Rat2 Embryonic Fibroblasts. J Toxicol. , 323828 (2010).
  10. Besaratinia, A., Bates, S. E., Pfeifer, G. P. Mutational signature of the proximate bladder carcinogen N-hydroxy-4-acetylaminobiphenyl: inconsistency with the p53 mutational spectrum in bladder cancer. Cancer Res. 62 (15), 4331-4338 (2002).
  11. Besaratinia, A., Pfeifer, G. P. Enhancement of the mutagenicity of benzo(a)pyrene diol epoxide by a nonmutagenic dose of ultraviolet A radiation. Cancer Res. 63 (24), 8708-8716 (2003).
  12. Besaratinia, A., Pfeifer, G. P. Weak yet distinct mutagenicity of acrylamide in mammalian cells. J Natl Cancer Inst. 95 (12), 889-896 (2003).
  13. Besaratinia, A., Pfeifer, G. P. Biological consequences of 8-methoxypsoralen-photoinduced lesions: sequence-specificity of mutations and preponderance of T to C and T to a mutations. J Invest Dermatol. 123 (6), 1140-1146 (2004).
  14. Besaratinia, A., Pfeifer, G. P. Genotoxicity of acrylamide and glycidamide. J Natl Cancer Inst. 96 (13), 1023-1029 (2004).
  15. Besaratinia, A., Bates, S. E., Synold, T. W., Pfeifer, G. P. Similar mutagenicity of photoactivated porphyrins and ultraviolet A radiation in mouse embryonic fibroblasts: involvement of oxidative DNA lesions in mutagenesis. Biochemistry. 43 (49), 15557-15566 (2004).
  16. Yoon, J. H., et al. DNA damage, repair, and mutation induction by (+)-Syn and (-)-anti-dibenzo[a,l]pyrene-11,12-diol-13,14-epoxides in mouse cells. Cancer Res. 64 (20), 7321-7328 (2004).
  17. Besaratinia, A., Synold, T. W., Xi, B., Pfeifer, G. P. G-to-T transversions and small tandem base deletions are the hallmark of mutations induced by ultraviolet a radiation in mammalian cells. Biochemistry. 43 (25), 8169-8177 (2004).
  18. Besaratinia, A., Pfeifer, G. P. Investigating DNA adduct-targeted mutagenicity of tamoxifen: preferential formation of tamoxifen-DNA adducts in the human p53 gene in SV40 immortalized hepatocytes but not endometrial carcinoma cells. Biochemistry. 44 (23), 8418-8427 (2005).
  19. Kim, S. I., Pfeifer, G. P., Besaratinia, A. Lack of mutagenicity of acrolein-induced DNA adducts in mouse and human cells. Cancer Res. 67 (24), 11640-11647 (2007).
  20. Besaratinia, A., Kim, S. I., Bates, S. E., Pfeifer, G. P. Riboflavin activated by ultraviolet A1 irradiation induces oxidative DNA damage-mediated mutations inhibited by vitamin C. Proc Natl Acad Sci U S A. 104 (14), 5953-5958 (2007).
  21. Besaratinia, A., Kim, S. I., Pfeifer, G. P. Rapid repair of UVA-induced oxidized purines and persistence of UVB-induced dipyrimidine lesions determine the mutagenicity of sunlight in mouse cells. FASEB J. 22 (7), 2379-2392 (2008).
  22. Besaratinia, A., Kim, S. I., Hainaut, P., Pfeifer, G. P. In vitro recapitulating of TP53 mutagenesis in hepatocellular carcinoma associated with dietary aflatoxin B1 exposure. Gastroenterology. 137 (3), (2009).
  23. Besaratinia, A., et al. A high-throughput next-generation sequencing-based method for detecting the mutational fingerprint of carcinogens. Nucleic Acids Res. 40 (15), e116 (2012).
  24. Tommasi, S., Bates, S. E., Behar, R. Z., Talbot, P., Besaratinia, A. Limited mutagenicity of electronic cigarettes in mouse or human cells in vitro. Lung Cancer. 112, 41-46 (2017).
  25. Dycaico, M. J., et al. The use of shuttle vectors for mutation analysis in transgenic mice and rats. Mutat Res. 307 (2), 461-478 (1994).
  26. Davies, R., et al. Mutational spectra of tamoxifen-induced mutations in the livers of lacI transgenic rats. Environ Mol Mutagen. 28 (4), 430-433 (1996).
  27. de Boer, J. G., et al. Spectrum of spontaneous mutations in liver tissue of lacI transgenic mice. Environ Mol Mutagen. 30 (3), 273-286 (1997).
  28. de Boer, J. G., Mirsalis, J. C., Provost, G. S., Tindall, K. R., Glickman, B. W. Spectrum of mutations in kidney, stomach, and liver from lacI transgenic mice recovered after treatment with tris(2,3-dibromopropyl)phosphate. Environ Mol Mutagen. 28 (4), 418-423 (1996).
  29. Dycaico, M. J., et al. Species-specific differences in hepatic mutant frequency and mutational spectrum among lambda/lacI transgenic rats and mice following exposure to aflatoxin B1. Carcinogenesis. 17 (11), 2347-2356 (1996).
  30. Skopek, T. R., Kort, K. L., Marino, D. R. Relative sensitivity of the endogenous hprt gene and lacI transgene in ENU-treated Big Blue B6C3F1 mice. Environ Mol Mutagen. 26 (1), 9-15 (1995).
  31. Skopek, T. R., et al. Mutagenic response of the endogenous hprt gene and lacI transgene in benzo[a]pyrene-treated Big Blue B6C3F1 mice. Environ Mol Mutagen. 28 (4), 376-384 (1996).
  32. Erfle, H. L., et al. An efficient laboratory protocol for the sequencing of large numbers of lacI mutants recovered from Big Blue transgenic animals. Environ Mol Mutagen. 28 (4), 393-396 (1996).
  33. Gu, M., Ahmed, A., Wei, C., Gorelick, N., Glickman, B. W. Development of a lambda-based complementation assay for the preliminary localization of lacI mutants from the Big Blue mouse: implications for a DNA-sequencing strategy. Mutat Res. 307 (2), 533-540 (1994).
  34. Harbach, P. R., Zimmer, D. M., Filipunas, A. L., Mattes, W. B., Aaron, C. S. Spontaneous mutation spectrum at the lambda cII locus in liver, lung, and spleen tissue of Big Blue transgenic mice. Environ Mol Mutagen. 33 (2), 132-143 (1999).
  35. Kohler, S. W., et al. Spectra of spontaneous and mutagen-induced mutations in the lacI gene in transgenic mice. Proc Natl Acad Sci U S A. 88 (18), 7958-7962 (1991).
  36. Mittelstaedt, R. A., et al. Comparison of the types of mutations induced by 7,12-dimethylbenz[a]anthracene in the lacI and hprt genes of Big Blue rats. Environ Mol Mutagen. 31 (2), 149-156 (1998).
  37. Monroe, J. J., Kort, K. L., Miller, J. E., Marino, D. R., Skopek, T. R. A comparative study of in vivo mutation assays: analysis of hprt, lacI, cII/cI and as mutational targets for N-nitroso-N-methylurea and benzo[a]pyrene in Big Blue mice. Mutat Res. 421 (1), 121-136 (1998).
  38. Morrison, V., Ashby, J. A preliminary evaluation of the performance of the Muta Mouse (lacZ) and Big Blue (lacI) transgenic mouse mutation assays. Mutagenesis. 9 (4), 367-375 (1994).
  39. Okonogi, H., et al. Agreement of mutational characteristics of heterocyclic amines in lacI of the Big Blue mouse with those in tumor related genes in rodents. Carcinogenesis. 18 (4), 745-748 (1997).
  40. Provost, G. S., Mirsalis, J. C., Rogers, B. J., Short, J. M. Mutagenic response to benzene and tris(2,3-dibromopropyl)-phosphate in the lambda lacI transgenic mouse mutation assay: a standardized approach to in vivo mutation analysis. Environ Mol Mutagen. 28 (4), 342-347 (1996).
  41. Shane, B. S., et al. LacI mutation spectra following benzo[a]pyrene treatment of Big Blue mice. Carcinogenesis. 21 (4), 715-725 (2000).
  42. Shane, B. S., Lockhart, A. M., Winston, G. W., Tindall, K. R. Mutant frequency of lacI in transgenic mice following benzo[a]pyrene treatment and partial hepatectomy. Mutat Res. 377 (1), 1-11 (1997).
  43. Shephard, S. E., Sengstag, C., Lutz, W. K., Schlatter, C. Mutations in liver DNA of lacI transgenic mice (Big Blue) following subchronic exposure to 2-acetylaminofluorene. Mutat Res. 302 (2), 91-96 (1993).
  44. Stiegler, G. L., Stillwell, L. C. Big Blue transgenic mouse lacI mutation analysis. Environ Mol Mutagen. 22 (3), 127-129 (1993).
  45. Walker, V. E., et al. Frequency and spectrum of ethylnitrosourea-induced mutation at the hprt and lacI loci in splenic lymphocytes of exposed lacI transgenic mice. Cancer Res. 56 (20), 4654-4661 (1996).
  46. Young, R. R., Rogers, B. J., Provost, G. S., Short, J. M., Putman, D. L. Interlaboratory comparison: liver spontaneous mutant frequency from lambda/lacI transgenic mice (Big Blue) (II). Mutat Res. 327 (1-2), 67-73 (1995).
  47. Zimmer, D. M., Zhang, X. B., Harbach, P. R., Mayo, J. K., Aaron, C. S. Spontaneous and ethylnitrosourea-induced mutation fixation and molecular spectra at the lacI transgene in the Big Blue rat-2 embryo cell line. Environ Mol Mutagen. 28 (4), 325-333 (1996).
  48. Swiger, R. R., et al. The cII locus in the MutaMouse system. Environ Mol Mutagen. 34 (2-3), 201-207 (1999).
  49. Heddle, J. A., Martus, H. J., Douglas, G. R. Treatment and sampling protocols for transgenic mutation assays. Environ Mol Mutagen. 41 (1), 1-6 (2003).
  50. Thybaud, V., et al. In vivo transgenic mutation assays. Mutat Res. 540 (2), 141-151 (2003).
  51. Manjanatha, M. G., Cao, X., Shelton, S. D., Mittelstaedt, R. A., Heflich, R. H. In vivo cII, gpt, and Spi(-) gene mutation assays in transgenic mice and rats. Methods Mol Biol. 1044, 97-119 (2013).
  52. Swiger, R. R. Quantifying in vivo somatic mutations using transgenic mouse model systems. Methods Mol Biol. 1105, 271-282 (2014).
  53. Tommasi, S., Besaratinia, A., Wilczynski, S. P., Pfeifer, G. P. Loss of Rassf1a enhances p53-mediated tumor predisposition and accelerates progression to aneuploidy. Oncogene. 30 (6), 690-700 (2011).
  54. Saluz, H. P., Jost, J. P. . A Laboratory Guide to Genomic Sequencing. , (1987).
  55. Wijnholds, J., Philipsen, J. N., Ab, G. Tissue-specific and steroid-dependent interaction of transcription factors with the oestrogen-inducible apoVLDL II promoter in vivo. EMBO J. 7 (9), 2757-2763 (1988).
  56. . λ Select-cII Mutation Detection System for Big Blue Rodents. INSTRUCTION MANUAL; Catalog #720120; Revision B.0 Available from: https://www.agilent.com/cs/library/usermanuals/public/720120.pdf (2018)
  57. Adams, W. T., Skopek, T. R. Statistical test for the comparison of samples from mutational spectra. J Mol Biol. 194 (3), 391-396 (1987).
  58. Kim, S. I., Yoon, J. I., Tommasi, S., Besaratinia, A. New experimental data linking secondhand smoke exposure to lung cancer in nonsmokers. FASEB J. 26 (5), 1845-1854 (2012).
  59. Yoon, J. I., Kim, S. I., Tommasi, S., Besaratinia, A. Organ specificity of the bladder carcinogen 4-aminobiphenyl in inducing DNA damage and mutation in mice. Cancer Prev Res (Phila). 5 (2), 299-308 (2012).
  60. Boyiri, T., et al. Mammary carcinogenesis and molecular analysis of in vivo cII gene mutations in the mammary tissue of female transgenic rats treated with the environmental pollutant 6-nitrochrysene. Carcinogenesis. 25 (4), 637-643 (2004).
  61. Chen, T., et al. Mutations induced by alpha-hydroxytamoxifen in the lacI and cII genes of Big Blue transgenic rats. Carcinogenesis. 23 (10), 1751-1757 (2002).
  62. Chen, T., et al. 4-Aminobiphenyl induces liver DNA adducts in both neonatal and adult mice but induces liver mutations only in neonatal mice. Int J Cancer. 117 (2), 182-187 (2005).
  63. Crabbe, R. A., Hill, K. A. Heart tissue of harlequin (hq)/Big Blue mice has elevated reactive oxygen species without significant impact on the frequency and nature of point mutations in nuclear DNA. Mutat Res. 691 (1-2), 64-71 (2010).
  64. Hernandez, L. G., Heddle, J. A. A carcinogenic western diet does not induce somatic mutations in various target tissues of transgenic C56BL/6 mice. Mutat Res. 570 (2), 185-196 (2005).
  65. Manjanatha, M. G., et al. Dose and temporal evaluation of ethylene oxide-induced mutagenicity in the lungs of male big blue mice following inhalation exposure to carcinogenic concentrations. Environ Mol Mutagen. 58 (3), 122-134 (2017).
  66. Manjanatha, M. G., et al. Evaluation of mutagenic mode of action in Big Blue mice fed methylphenidate for 24 weeks. Mutat Res. 680 (1-2), 43-48 (2009).
  67. McDaniel, L. P., et al. Mutagenicity and DNA adduct formation by aristolochic acid in the spleen of Big Blue(R) rats. Environ Mol Mutagen. 53 (5), 358-368 (2012).
  68. Mei, N., Heflich, R. H., Moore, M. M., Chen, T. Age-dependent sensitivity of Big Blue transgenic mice to the mutagenicity of N-ethyl-N-nitrosourea (ENU) in liver. Mutat Res. 572 (1-2), 14-26 (2005).
  69. Mei, N., et al. The genotoxicity of acrylamide and glycidamide in big blue rats. Toxicol Sci. 115 (2), 412-421 (2010).
  70. Nay, S. L., Lee, D. H., Bates, S. E., O’Connor, T. R. Alkbh2 protects against lethality and mutation in primary mouse embryonic fibroblasts. DNA Repair (Amst). 11 (5), 502-510 (2012).
  71. Singh, V. K., Ganesh, L., Cunningham, M. L., Shane, B. S. Comparison of the mutant frequencies and mutation spectra of three non-genotoxic carcinogens, oxazepam, phenobarbital, and Wyeth 14,643, at the lambdacII locus in Big Blue transgenic mice. Biochem Pharmacol. 62 (6), 685-692 (2001).
  72. Stuart, G. R., et al. Interpretation of mutational spectra from different genes: analyses of PhIP-induced mutational specificity in the lacI and cII transgenes from colon of Big Blue rats. Mutat Res. 452 (1), 101-121 (2000).
  73. Terrell, A. N., et al. Mutagenicity of furan in female Big Blue B6C3F1 mice. Mutat Res Genet Toxicol Environ Mutagen. 770, 46-54 (2014).
  74. Thompson, C. M., et al. Assessment of the mutagenic potential of Cr(VI) in the oral mucosa of Big Blue(R) transgenic F344 rats. Environ Mol Mutagen. 56 (7), 621-628 (2015).
  75. Wang, J., Liu, X., Heflich, R. H., Chen, T. Time course of cII gene mutant manifestation in the liver, spleen, and bone marrow of N-ethyl-N-nitrosourea-treated Big Blue transgenic mice. Toxicol Sci. 82 (1), 124-128 (2004).
  76. LeBlanc, G. A., Bain, L. J. Chronic toxicity of environmental contaminants: sentinels and biomarkers. Environ Health Perspect. 105, 65-80 (1997).
  77. Arlt, V. M., et al. Aristolochic acid mutagenesis: molecular clues to the aetiology of Balkan endemic nephropathy-associated urothelial cancer. Carcinogenesis. 28 (11), 2253-2261 (2007).
  78. Besaratinia, A., Pfeifer, G. P. Second-hand smoke and human lung cancer. Lancet Oncol. 9 (7), 657-666 (2008).

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Lambda Select CIIMutation DetectionTransgenic RodentsMutagenicity TestingMammalian CellsReporter GenePackaging ReactionSM BufferChloroformPlating CultureTiteringScreeningE. ColiPhagesTB1 Top Agar

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Besaratinia, A., Tommasi, S. The Lambda Select cII Mutation Detection System. J. Vis. Exp. (134), e57510, doi:10.3791/57510 (2018).
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