Establishment of Qualitative PCR Detection of Transgenic Insect⁃resistant Maize CM8101

doi:10.19586/j.2095-2341.2021.0158

Current Biotechnology ›› 2022, Vol. 12 ›› Issue (2): 248-255.DOI: 10.19586/j.2095-2341.2021.0158

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Establishment of Qualitative PCR Detection of Transgenic Insect⁃resistant Maize CM8101

Hongtao WEN¹(), Yang YANG¹(), Yijia DING¹, Ran YUAN¹, Ruiying ZHANG¹, Zhiyuan XU²(), Jingang LIANG³()

^1.Quality & Safety Institute of Agricultural Products，Heilongjiang Academy of Agricultural Sciences，Harbin 150086，China
^2.College of Agriculture，Northeast Agricultural University，Harbin 150038，China
^3.Development Center of Science and Technology，Ministry of Agriculture and Rural Affairs，Beijing 100176，China

Received:2021-09-23 Accepted:2021-10-22 Online:2022-03-25 Published:2022-03-25
Contact: Zhiyuan XU,Jingang LIANG

抗虫玉米CM8101定性检测方法的建立

温洪涛¹(), 杨洋¹(), 丁一佳¹, 苑冉¹, 张瑞英¹, 徐志远²(), 梁晋刚³()

^1.黑龙江省农业科学院农产品质量安全研究所，哈尔滨 150086
^2.东北农业大学农学院，哈尔滨 150038
^3.农业农村部科技发展中心，北京 100176

通讯作者: 徐志远,梁晋刚
作者简介:温洪涛与杨洋为本文共同第一作者。温洪涛E-mail：wen0891@163.com
杨洋E-mail：yangy_yx@163.com
基金资助:
国家转基因新品种培育重大专项(2018ZX08011-04B)

Abstract

Abstract:

Transgenic insect-resistant maize CM8101 is a new strain with modified Bt gene Cry1Ab?Ma syntheszied by artificial transformation. It is developed by China and has better insect resistance. At present， this strain has entered the stage of productive test， and it has important industrial application prospect in China. According to the 5′ flanking sequence information of transgenic maize CM8101， the optimal specific primer and probe combinations were designed and screened. A normal PCR assay and a real-time PCR assay for transgenic insect-resistant maize CM8101 were developed in this paper. Both methods could be used to qualitatively detect the component of transgenic insect-resistant maize CM8101 with good specificity. The results showed that the detection limit of normal PCR method was up to 0.1%， and the detection limit of real-time PCR method was up to 0.05%. Establishment of the two qualitative PCR methods provides new reference method for accurate and efficient detection of transgenic insect resistant maize CM8101 and its products in the future.

Key words: genetically modified maize, CM8101 event, qualitative detection

摘要：

转基因抗虫玉米CM8101是经人工改造合成的转Bt抗虫基因Cry1Ab?Ma转基因玉米新品系，由我国自主研发，具有更优良的抗虫性，目前已进入生产性试验阶段，具有广阔的产业化应用前景。依据CM8101的5′端旁侧序列信息，设计并筛选出最佳引物探针组合，通过普通PCR和实时荧光PCR技术，建立了转基因抗虫玉米CM8101的两种特异性定性检测方法。结果显示，普通PCR检测方法检出限达0.1%，实时荧光PCR检测方法检出限达0.05%。这两种定性检测方法的建立为今后准确高效检测转基因抗虫玉米CM8101及其产品提供了新的参考方法。

关键词: 转基因玉米, CM8101转化体, 定性检测

CLC Number:

Q785

Hongtao WEN, Yang YANG, Yijia DING, Ran YUAN, Ruiying ZHANG, Zhiyuan XU, Jingang LIANG. Establishment of Qualitative PCR Detection of Transgenic Insect⁃resistant Maize CM8101[J]. Current Biotechnology, 2022, 12(2): 248-255.

温洪涛, 杨洋, 丁一佳, 苑冉, 张瑞英, 徐志远, 梁晋刚. 抗虫玉米CM8101定性检测方法的建立[J]. 生物技术进展, 2022, 12(2): 248-255.

Figures/Tables 12

References 20

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引物/探针	序列(5′→3′)	引物/探针	序列(5′→3′)
CM8101⁃LB⁃F1	5′⁃AGCAAGTGTCGAGTTCAGCTCT⁃3′	CM8101⁃LB⁃R1	5′⁃AGCAATTCGGCGTTAATTCAGT⁃3′
CM8101⁃LB⁃F2	5′⁃AGCAATTAAGCAAGTGTCGAGT⁃3′	CM8101⁃LB⁃R2	5′⁃GGCGAATGCTAGAGCAATTCG⁃3′
CM8101⁃LB⁃F3	5′⁃TCAACCCATGTTAGTCTTAGCA⁃3′	CM8101⁃LB⁃R3	5′⁃CAGCCTGAATGGCGAATGCTA⁃3′
CM8101⁃LB-F4	5′⁃CGGTGCCAGAGTCAACCCAT⁃3′	CM8101⁃LB⁃R4	5′⁃CTTCCCAACAGTTGCGCAG⁃3′
CM8101⁃LB⁃F5	5′⁃TCATCCCAACATGAGCTAGTCG⁃3′	CM8101⁃LB⁃R5	5′⁃ACCCTGGCGTTACCCAACTT⁃3′
CM8101⁃LB⁃F6	5′⁃AGTCCTAAAATGTGTTCTCCCT⁃3′	CM8101⁃LB⁃qP	5′⁃FAM⁃AGCTGACCCTTCTAACACATTGCGGACGTT⁃BHQ1⁃3′

引物/探针	序列(5′→3′)	引物/探针	序列(5′→3′)
CM8101⁃LB⁃F1	5′⁃AGCAAGTGTCGAGTTCAGCTCT⁃3′	CM8101⁃LB⁃R1	5′⁃AGCAATTCGGCGTTAATTCAGT⁃3′
CM8101⁃LB⁃F2	5′⁃AGCAATTAAGCAAGTGTCGAGT⁃3′	CM8101⁃LB⁃R2	5′⁃GGCGAATGCTAGAGCAATTCG⁃3′
CM8101⁃LB⁃F3	5′⁃TCAACCCATGTTAGTCTTAGCA⁃3′	CM8101⁃LB⁃R3	5′⁃CAGCCTGAATGGCGAATGCTA⁃3′
CM8101⁃LB-F4	5′⁃CGGTGCCAGAGTCAACCCAT⁃3′	CM8101⁃LB⁃R4	5′⁃CTTCCCAACAGTTGCGCAG⁃3′
CM8101⁃LB⁃F5	5′⁃TCATCCCAACATGAGCTAGTCG⁃3′	CM8101⁃LB⁃R5	5′⁃ACCCTGGCGTTACCCAACTT⁃3′
CM8101⁃LB⁃F6	5′⁃AGTCCTAAAATGTGTTCTCCCT⁃3′	CM8101⁃LB⁃qP	5′⁃FAM⁃AGCTGACCCTTCTAACACATTGCGGACGTT⁃BHQ1⁃3′

序号	终浓度/(μmol·L^-1)		序号	终浓度/(μmol·L^-1)
序号	引物	探针	序号	引物	探针
1	0.2	0.2	9	0.4	0.2
2		0.3	10		0.3
3		0.4	11		0.4
4		0.5	12		0.5
5	0.3	0.2	13	0.5	0.2
6		0.3	14		0.3
7		0.4	15		0.4
8		0.5	16		0.5

序号	终浓度/(μmol·L^-1)		序号	终浓度/(μmol·L^-1)
序号	引物	探针	序号	引物	探针
1	0.2	0.2	9	0.4	0.2
2		0.3	10		0.3
3		0.4	11		0.4
4		0.5	12		0.5
5	0.3	0.2	13	0.5	0.2
6		0.3	14		0.3
7		0.4	15		0.4
8		0.5	16		0.5

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Establishment of Qualitative PCR Detection of Transgenic Insect⁃resistant Maize CM8101

抗虫玉米CM8101定性检测方法的建立

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