This paper discusses the modified CTAB method for DNA extraction of toxigenic Aspergillus and compares it with the kit extraction method. The modified CTAB method successfully extracted DNA of toxigenic Aspergillus. The concentration and purity reached the standard of PCR amplification. Two pairs of primers were designed based on the aflR gene for aflatoxin biosynthesis regulation, and the toxigenic Aspergillus aflR gene was identified by using PCR amplification. The results show that the aflR gene is contained in the genome of Aspergillus flavus, Aspergillus parasitic and Aspergillus tamarii, Aspergillus fumigates and Aspergillus versicolor are not detected, realizing the purpose for identifying aflatoxin producing strains. This method provides a reference for rapid detection of toxigenic fungi.
JIANG Dan
,
JIN Weilin
,
SU Deshan
,
HUANG Yaojiang
. DNA Preparation of Toxigenic Aspergillus and PCR Detection of aflR Toxin Gene[J]. Science & Technology Review, 2014
, 32(14)
: 31
-34
.
DOI: 10.3981/j.issn.1000-7857.2014.14.004
[1] 李群伟, 王绍萍, 鲍文生. 真菌毒素与人类疾病的研究与展望[J]. 中国 地方病防治杂志, 2001, 16(1): 24-25. Li Qunwei, Wang Shaoping, Bao Wensheng. The research progress and prospect of mycotoxin and human disease[J]. Chinese Journal of Control of Endemic Disenaces, 2001, 16(1): 24-25.
[2] 邢来君, 李明春. 普通真菌学[M]. 北京: 高等教育出版社, 1999: 11-22. Xing Laijun, Li Mingchun. Ordinary mycology[M]. Beijing: Higher Education Press, 1999: 11-22.
[3] 李鹏, 赖卫华, 金晶. 食品中真菌毒素的研究[J]. 农产品加工·学刊, 2005(3): 12-15. Li Peng, Lai Weihua, Jin Jing. Progress of studying on mycotoxins in food[J]. Academic Periodical of Farm Products Processing, 2005(3): 12-15.
[4] 齐祖同. 中国真菌志[M]. 5卷. 北京: 科学出版社, 1997: 1-4. Qi Zutong. Flora fungorum sinicorum[M]. Vol 5. Beijing: Science Press, 1997: 1-4.
[5] Brera C, Miraglia M, Colatosti M. Evaluation of the impact of mycotoxins on human health: Sources of errors[J]. Microchemical Journal, 1998, 59 (1): 45-49.
[6] Papp E, H-Otta K, Záray Z, et al. Liquid chromatographic determination of aflatoxins[J]. Microchemical Journal, 2002, 73(1): 39-46.
[7] 苏福荣, 王松雪, 孙辉, 等. 国内外粮食中真菌毒素限量标准制定的现 状与分析[J]. 粮油食品科技, 2007, 15(6): 57-59. Su Furong, Wang Songxue, Sun Hui, et al. The state of mycotoxin maximum limit of grain[J]. Science and Technology of Cereals, Oils and Foods, 2007, 15(6): 57-59.
[8] 谷艳昌, 侯瑛, 王海磊, 等. 一种简易有效的提取真菌DNA化学改良 方法[J]. 安徽农业科学, 2009, 37(15): 6884-6886. Gu Yanchang, Hou Ying, Wang Hailei, et al. A simple and effective chemical modification method for extracting DNA from fungi[J]. Journal of Anhui Agricultural Sciences, 2009, 37(15): 6884-6886.
[9] 周小玲, 沈微, 饶志明, 等. 一种快速提取真菌染色体DNA的方法[J]. 微生物学通报, 2004, 31(4): 89-92. Zhou Xiaoling, Shen Wei, Rao Zhiming, et al. A rapid method for preparation of fungal chromosome DNA[J]. Microbiology, 2004, 31(4): 89-92.
[10] Yu J J, Chang P K, Ehrlich K C, et al. Clustered pathway genes in aflatoxin biosynthesis[J]. Applied and Environmental Microbiology, 2004, 70(3): 1253-1262.
[11] Somashekar D, Rati E R, Chandrashekar A. PCR-restriction fragment length analysis of aflR gene for differentiation and detection of Aspergillus flavus and Aspergillus parasiticus in maize[J]. International Journal of Food Microbiology, 2004, 93(1): 101-107.
[12] 吴发红, 黄东益, 黄小龙, 等. 几种真菌DNA提取方法的比较[J]. 中 国农学通报, 2009, 25(8): 62-64. Wu Fahong, Huang Dongyi, Huang Xiaolong, et al. Comparing study on several methods for DNA extraction from endophytic fungi[J]. Chinese Agricultural Science Bulletin, 2009, 25(8): 62-64.
[13] Cheng F S, Brown S K, Weeden N F. A DNA extraction protocol from various tissues in woody species[J]. Hortscience, 1997, 32(5): 921-922.
[14] 陈桂信, 吕柳新, 赖钟雄, 等. 基因组DNA的提取与纯化[J]. 江西农 业大学学报, 2004, 26(3): 329-331. Chen Guixin, Lü liuxin, Lai Zhongxiong, et al. Extraction and purification of genomic DNA in Nai (Prunus salicina Lindl. var. cordata)[J]. Acta Agriculturae Universitatis Jiangxiensis, 2004, 26(3): 329-331.
[15] Goto T, Wicklow D T, Ito Y. Aflatoxin and cyclopiazonic acid production by a sclerotium-producing Aspergillus tamarii strain [J]. Applied and Environmental Microbiology, 1996, 62(11): 4036-4038.
