A Qualitative Detection Method for Genetically Modified Maize KJ1003 by Real-time PCR

doi:10.19586/j.2095-2341.2026.0027

Current Biotechnology ›› 2026, Vol. 16 ›› Issue (4): 833-842.DOI: 10.19586/j.2095-2341.2026.0027

• Special Forum on Detection Technology for Genetically Modified Organisms • Previous Articles Next Articles

A Qualitative Detection Method for Genetically Modified Maize KJ1003 by Real-time PCR

Tongtong ZHAO(), Ziyan CHEN, Hua ZHANG, Xinyao PEI, Chenyao WANG, Jiake LI, Dan WANG, Yueqiu ZHANG, Wei FU(), Hong CHEN()

Development Center of Science and Technology，Ministry of Agriculture and Rural Affairs，Beijing 100176，China

Received:2026-02-06 Accepted:2026-04-03 Online:2026-07-25 Published:2026-09-11
Contact: Wei FU,Hong CHEN

Abstract

Abstract:

The genetically modified maize line KJ1003 is a transformation event developed by Kejidalong （Beijing） Biotechnology Co.， Ltd. It exhibits excellent insect resistance and herbicide tolerance， has obtained a genetically modified organism safety certificate， and possesses promising industrialization prospects. In this study， a real-time fluorescent PCR qualitative detection method for the KJ1003 transformation event was established， targeting the specific sequence of KJ1003 through primer and probe design and screening， reaction system optimization， as well as specificity， sensitivity， limit of detection， and robustness tests， together with inter-laboratory validation. The results showed that this method could effectively detect KJ1003 transgenic components and amplify a 158 bp specific fragment， with a detection sensitivity and limit of detection both reaching 0.05% and favorable robustness. The verification results of the method′s specificity， limit of detection and reproducibility conducted by 8 laboratories all complied with relevant standard requirements. This study provides a scientific basis and technical support for the detection and safety supervision of genetically modified crops in China.

Key words: genetically modified maize, insect-tolerant and herbicide-tolerant, KJ1003, real-time PCR, qualitative detection

CLC Number:

Q789

Tongtong ZHAO, Ziyan CHEN, Hua ZHANG, Xinyao PEI, Chenyao WANG, Jiake LI, Dan WANG, Yueqiu ZHANG, Wei FU, Hong CHEN. A Qualitative Detection Method for Genetically Modified Maize KJ1003 by Real-time PCR[J]. Current Biotechnology, 2026, 16(4): 833-842.

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Figures/Tables 11

References 27

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转基因混合样品	组成成分（每种转化体质量分数1%）
其他转基因玉米混合样	Bt11、Bt176、MON810、MON863、GA21、NK603、T25、TC1507、MON89034、MON88017、59122、MIR604、3272、MON87460、DAS40278-9、4114、MON87427、5307、DBN9858、ND207、DBN9501、瑞丰125、DBN9936、MON87411、MZIR098、浙大瑞丰8、BFL4-2、nCX-1
转基因大豆混合样	GTS40-3-2、MON89788、A5547-127、A2704-12、356043、305423、CV127、MON87701、MON87708、MON87769、MON87705、FG72、DAS81419-2、DAS-444Ø6-6、MON87751、SHZD3201、CAL16、中黄6106、DBN9004
转基因水稻混合样	TT51-1、KF-6、KMD-1、M12、KF-8、KF-2、G6H1、T1C-19
转基因油菜混合样	MS1、MS8、RF1、RF2、RF3、T45、Oxy-235、Topas19/2、MON88302、73496
转基因棉花混合样	MON1445、MON531、MON15985、LLCOTTON25、MON88913、GHB614、COT102
非转基因玉米混合样	多种非转基因玉米等量混合制成

转基因混合样品	组成成分（每种转化体质量分数1%）
其他转基因玉米混合样	Bt11、Bt176、MON810、MON863、GA21、NK603、T25、TC1507、MON89034、MON88017、59122、MIR604、3272、MON87460、DAS40278-9、4114、MON87427、5307、DBN9858、ND207、DBN9501、瑞丰125、DBN9936、MON87411、MZIR098、浙大瑞丰8、BFL4-2、nCX-1
转基因大豆混合样	GTS40-3-2、MON89788、A5547-127、A2704-12、356043、305423、CV127、MON87701、MON87708、MON87769、MON87705、FG72、DAS81419-2、DAS-444Ø6-6、MON87751、SHZD3201、CAL16、中黄6106、DBN9004
转基因水稻混合样	TT51-1、KF-6、KMD-1、M12、KF-8、KF-2、G6H1、T1C-19
转基因油菜混合样	MS1、MS8、RF1、RF2、RF3、T45、Oxy-235、Topas19/2、MON88302、73496
转基因棉花混合样	MON1445、MON531、MON15985、LLCOTTON25、MON88913、GHB614、COT102
非转基因玉米混合样	多种非转基因玉米等量混合制成

引物和探针（5′端）	序列（5′→3′）	引物和探针（3′端）	序列（5′→3′）
RB-qF1	5′-CGTGTCCTCGCTCCCTTTTC-3′	LB-qF1	5′-CGCGTGAATACGTAGCTGTG-3′
RB-qF2	5′-GCTCCCTTTTCTCTTCCCCAA-3′	LB-qF2	5′-TGTGAAAAGAGAACGCGTGAAT-3′
RB-qF3	5′-GTCCTCGCTCCCTTTTCTCTT-3′	LB-qF3	5′-TGTGTCCCAAGTAAACCTCAGT-3′
RB-qR1	5′-ATAGCGCGCAAACTAGGATAAA-3′	LB-qR1	5′-CCCAAGTCCCACTCCTTGTG-3′
RB-qR2	5′-AGCGCGCAAACTAGGATAA-3′	LB-qR2	5′-AACCGGACCGTGGTGTTTT-3′
RB-qR3	5′-ATAGCGCGCAAACTAGGAT-3′	LB-qR3	5′-GCGCGTAAGTCTTGGTCTTC-3′
RB-qP1	5′-FAM-CAGATTGTCGTTTCCCGCCT-BHQ1-3′	LB-qP1	5′-FAM-GTGGCTCTAGCCGCTCCCAG-BHQ1-3′
RB-qP2	5′-FAM-TGTCGTTTCCCGCCTTCAGT-BHQ1-3′	LB-qP2	5′-FAM-CTCTAGCCGCTCCCAGTCGA-BHQ1-3′

引物和探针（5′端）	序列（5′→3′）	引物和探针（3′端）	序列（5′→3′）
RB-qF1	5′-CGTGTCCTCGCTCCCTTTTC-3′	LB-qF1	5′-CGCGTGAATACGTAGCTGTG-3′
RB-qF2	5′-GCTCCCTTTTCTCTTCCCCAA-3′	LB-qF2	5′-TGTGAAAAGAGAACGCGTGAAT-3′
RB-qF3	5′-GTCCTCGCTCCCTTTTCTCTT-3′	LB-qF3	5′-TGTGTCCCAAGTAAACCTCAGT-3′
RB-qR1	5′-ATAGCGCGCAAACTAGGATAAA-3′	LB-qR1	5′-CCCAAGTCCCACTCCTTGTG-3′
RB-qR2	5′-AGCGCGCAAACTAGGATAA-3′	LB-qR2	5′-AACCGGACCGTGGTGTTTT-3′
RB-qR3	5′-ATAGCGCGCAAACTAGGAT-3′	LB-qR3	5′-GCGCGTAAGTCTTGGTCTTC-3′
RB-qP1	5′-FAM-CAGATTGTCGTTTCCCGCCT-BHQ1-3′	LB-qP1	5′-FAM-GTGGCTCTAGCCGCTCCCAG-BHQ1-3′
RB-qP2	5′-FAM-TGTCGTTTCCCGCCTTCAGT-BHQ1-3′	LB-qP2	5′-FAM-CTCTAGCCGCTCCCAGTCGA-BHQ1-3′

编号	引物名称及序列（5′→3′）	产物大小/bp
组合1	RB-qF1/RB-qR1/RB-qP1	161
组合2	RB-qF1/RB-qR1/RB-qP2	161
组合3	RB-qF2/RB-qR2/RB-qP1	150
组合4	RB-qF2/RB-qR2/RB-qP2	150
组合5	RB-qF3/RB-qR3/RB-qP1	158
组合6	RB-qF3/RB-qR3/RB-qP2	158
组合7	RB-qF1/RB-qR2/RB-qP1	159
组合8	RB-qF1/RB-qR2/RB-qP2	159
组合9	LB-qF1/LB-qR1/LB-qP1	188
组合10	LB-qF1/LB-qR1/LB-qP2	188
组合11	LB-qF2/LB-qR2/LB-qP1	176
组合12	LB-qF2/LB-qR2/LB-qP2	200
组合13	LB-qF3/LB-qR3/LB-qP1	200
组合14	LB-qF3/LB-qR3/LB-qP2	200
组合15	LB-qF2/LB-qR1/LB-qP1	200
组合16	LB-qF2/LB-qR1/LB-qP2	200

A Qualitative Detection Method for Genetically Modified Maize KJ1003 by Real-time PCR

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Figures/Tables 11

References 27

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样品名称	8家实验室验证结果
样品名称	Lab1	Lab2	Lab3	Lab4	Lab5	Lab6	Lab7	Lab8
空白对照	-	-	-	-	-	-	-	-
阴性对照	-	-	-	-	-	-	-	-
阳性对照	+	+	+	+	+	+	+	+
1% KJ1003	+	+	+	+	+	+	+	+
其他转基因玉米混合样	-	-	-	-	-	-	-	-
转基因大豆混合样	-	-	-	-	-	-	-	-
转基因水稻混合样	-	-	-	-	-	-	-	-
转基因油菜混合样	-	-	-	-	-	-	-	-
转基因棉花混合样	-	-	-	-	-	-	-	-
非转基因玉米混合样	-	-	-	-	-	-	-	-
0.05%KJ1003（检出限）	+	+	+	+	+	+	+	+

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