Crops ›› 2017, Vol. 33 ›› Issue (6): 37-44.doi: 10.16035/j.issn.1001-7283.2017.06.007

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Determining Blind Samples of Transgenic Maize and Transgenic Soybean

Li Lina,Jin Longguo,Xie Chuanxiao,Liu Changlin   

  1. Institute of Crop Sciences,Chinese Academy of Agricultural Sciences,Beijing 100081,China
  • Received:2017-07-03 Revised:2017-10-27 Online:2017-12-15 Published:2018-08-26
  • Contact: Changlin Liu

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Abstract:

Genetically modified crops developed rapidly in the world. China imports a large amount of products of genetically modified crops every year, and has put billions of money in genetically modified crops research. China also has established regulations for genetically modified crops and their products. Simple and reliable methods for detecting transgenic ingredients are very urgent in China. In this study, according to the detection protocols of genetically modified crops published by the Ministry of Agriculture of the People’s Republic of China, blind samples of soybean (W160982 and W160984) and maize (W160983 and W160985) were detected for transgenic ingredients using the PCR technology. As a result, W160983, W160985, W160982, and W160984 are the standard sample MON89034, TC1507, GTS40-3-2, and BPS-CV127-9, respectively. The results of the blind samples were validated by sequence alignment of the PCR product. This study provided an experimental protocol for the detection of transgenic maize and soybean.

Key words: Transgenic maize, Transgenic soybean, Blind sample, Polymerase chain reaction

Table 1

PCR primers for detecting transgenic ingredients in maize and soybean"

靶标Target 名称Name 序列(5'→3')Sequence 产物大小Amplicon size (bp) 作物Crop 参考文献Reference
zSSIIb zSSIIb-F CTCCCAATCCTTTGACATCTGC 151 玉米 [9]
zSSIIb-R TCGATTTCTCTCTTGGTGACAGG
MON88017 MON88017-F TTGTCCTGAACCCCTAAAATCC 199 玉米 [10]
MON88017-R CCCGGACATGAAGCCATTTA
GA21 GA21-F CTTATCGTTATGCTATTTGCAACTT 112 玉米 [11]
GA21-R TGGCTCGCGATCCTCCTCGCGTTTC
MIR604 MIR604-F TCGCGCGCGGTGTCATCTATG 142 玉米 [12]
MIR604-R CGCGACACACCTCGTTAGTTAA
MON87460 MON87460-F ATCCACCTGTCAGCTCAAGTT 368
 玉米 [13]
MON87460-R GGTATGTATATAGTGGCGATG
MON89034 MON89034-F GCTGCTACTACTATCAAGCCAATA 207 玉米 [14]
MON89034-R TGCTTTCGCCTATAAATACGAC
MIR162 MIR162-F CCCGGGTCTAGACAATTCAGT 97 玉米 [15]
MIR162-R GCCCAGTAAAACAACTACCACAAG
MON810 MON810-F ACCTCGAGATTTACCTGATCCG 326 玉米 [16]
MON810-R TGCTGCAGGTGGTCTTACATC
MON863 MON863-F GCACTCAAAGACCTGGCGAATGA 411 玉米 [17]
MON863-R CCATCTTTGGGACCACTGTCG
Bt176 Bt176-F TTCAAGCACGGGAACTGGC 270 玉米 [18]
Bt176-R GAGCGAGAACACGAGAAGAGG
DAS-40278-9 DAS-40278-9-F CCATTCAGGAGACCTCGCTTG 238 玉米 [19]
DAS-40278-9-R CGAGCTTCAATCACTTTATGG
TC1507 TC1507-F CTTGTGGTGTTTGTGGCTCT 279 玉米 [20]
TC1507-R TGGCTCCTCCTTCGTATGT
3272 3272-F CTGGCCGATAAACTGACCAT 221 玉米 [21]
3272-R CCAAACGTAAAACGGCTTGT
59122 59122-F CGTCCGCAATGTGTTATTAAG 273 玉米 [22]
59122-R TGACCAAGTGTCCACTTGAC
NK603 NK603-F ATGAATGACCTCGAGTAATCTTGTTAA 108 玉米 [23]
NK603-R AAGAGATAACAGGATCCACTCAAACACT
Lectin Lec-1672F GGGTGAGGATAGGGTTCTCTG 210 大豆 [24]
Lec-1881R GCGATCGAGTAGTGAGAGTCG
GTS40-3-2 GTS40-3-2-F TTCAAACCCTTCAATTTAACCGAT 370 大豆 [25]
GTS40-3-2-R AAGGATAGTGGGATTGTGCGTC
MON89788 MON89788-F CTGCTCCACTCTTCCTTT 223 大豆 [26]
MON89788-R AGACTCTGTACCCTGACCT
MON87701 MON87701-MF GCACGCTTAGTGTGTGTGTCAAAC 150 大豆 [27]
靶标Target 名称Name 序列(5'→3') Sequence 产物大小Amplicon size (bp) 作物Crop 参考文献Reference
MON87701-MR GGATCCGTCGACCTGCAGTTAAC
MON87705 MON87705-F CGCCAAATCGTGAAGTTTCTCATCT 318 大豆 [28]
MON87705-R CAGTGATAACAACACCCTGAGTCT
MON87708 MON87708-F CCATCATACTCATTGCTGATCCA 233 大豆 [29]
MON87708-R AGCCAATCAATCTCAGAACTGTC
MON87769 MON87769-F CCGGACATGAAGCCATTTAC 298 大豆 [30]
MON87769-R TCCTTGGAGGTCGTCTCATT
BPS-CV127-9 CV127-F CCTTCGCCGTTTAGTGTATAGG 238 大豆 [31]
CV127-R AGCAGGTTCGTTTAAGGATGAA
305423 305423-F CGTCAGGAATAAAGGAAGTACAGTA 235 大豆 [32]
305423-R GCCCTAAAGGATGCGTATAGAGT
356043 356043-F CTTTTGCCCGAGGTCGTTAG 145 大豆 [33]
356043-R GCCCTTTGGTCTTCTGAGACTG

Fig.1

Gel electrophoresis detection of genome DNA extracted from transgenic maize and transgenic soybean"

Fig.2

Detection of transgenic ingredients in maize M: marker DL2000; B: blank control; N: negative control, the same below. Ⅰ: a-e. detections of MON8801, GA21, MIR604, MON87460, MON89034 transgenic ingredients; Ⅱ: a-e. detections of MIR162, MON810, MON863, Bt176, DAS40278-9 transgenic ingredients; Ⅲ: a-d. detections of TC1507, 3272, 59122, NK603 transgenic ingredients.1,3,5,7,9: W160985, 2,4,6,8,10: W160983"

Fig.3

Detections of transgenic ingredients in soybean Ⅰ: a-e.detections of GTS40-3-2, MON89788, MON87701, MON87705, MON87708 transgenic ingredients;Ⅱ: a-d.detections of MON87769, BPS-CV127-9, 305423, 356043 transgenic ingredients.1,3,5,7,9:W160982 samples, 2,4,6,8,10:W160984 samples"

Fig.4

Detections of transgenic ingredients and the internal reference gene zSSIIb in maize S: positive control (S1: TC1507, S2: MON89034), 1:W160985, 2:W160983.Ⅰ: maize endogenous gene (zSSIIb) amplification; Ⅱ: confirmation of target gene TC1507; Ⅲ: confirmation of target gene MON89034"

Fig.5

Detections of transgenic ingredients and the internal reference gene Lectin in soybean S: positive control (S1: GTS40-3-2, S2: BPS-CV127-9). 1. W160982, 2.W160984. Ⅰ: soybean endogenous gene (Lectin) amplification; Ⅱ: confirmation of target gene BPS-CV127-9; Ⅲ: confirmation of target gene GTS40-3-2"

Fig.6

Sequence comparison The box indicates the primer sequence.Ⅰ: the sequence comparison of MON89034 and W160983; Ⅱ: the sequence comparison of TC1507 and W160985; Ⅲ: the sequence comparison of GTS40-3-2 and W160982; Ⅳ: the sequence comparison of BPS-CV127-9 and W160984"

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