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Method for rapidly identifying specificity of transgenic herbicide-resistant soybean ZH10-6 transformant

A herbicide-tolerant and transgenic technology, which is applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., to achieve the effects of good specificity, improved detection efficiency, and easy operation.

Pending Publication Date: 2020-12-04
天津市农业科学院 +1
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Problems solved by technology

At present, there is no report on the detection method of the molecular characteristics of ZH10-6. In this study, the four border sequences of ZH10-6 were detected by multiplex PCR technology, and the identification method of the molecular characteristics of the transformant was established. Safety evaluation, administrative supervision and intellectual property protection provide important technical support

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  • Method for rapidly identifying specificity of transgenic herbicide-resistant soybean ZH10-6 transformant
  • Method for rapidly identifying specificity of transgenic herbicide-resistant soybean ZH10-6 transformant
  • Method for rapidly identifying specificity of transgenic herbicide-resistant soybean ZH10-6 transformant

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Embodiment 1

[0036] (1) Reagent: GO Taq produced by Promega Green Master Mix, 2×premix; endogenous soybean synthesized by Shanghai Sangong Lectin Gene and transformant specific sequence amplification primers. Includes ZH10A (Boundary A):

[0037] Primer forward sequence: 5'-GAGCCTCGGATGGGATTGAT-3' SEQ ID NO:1

[0038] Reverse primer sequence: 5'-TCGCTCATGTGTTGAGCATATAA-3' SEQ ID NO:2

[0039] ZH10B (boundary B):

[0040] Primer forward sequence: 5'-GTCGTGACTGGGAAAACCCT-3' SEQ ID NO:3

[0041] Reverse primer sequence: 5'- ATTCTAAAACGCTTTTCTTCTTGCTA-3' SEQ ID NO:4

[0042] ZH10C (boundary C):

[0043] Primer forward sequence: 5'-CTAAACCTCTTAATGATACGGCTT-3' SEQ ID NO:5

[0044] Reverse primer sequence: 5'- TCGTGACTGGGAAAACCCT-3' SEQ ID NO:6

[0045] ZH10D (boundary D)

[0046] Primer forward sequence: 5'- ACCCTTGTTGAACGAGTGG-3' SEQ ID NO:7

[0047] Reverse primer sequence: 5'- AGAAGAGTATTCGGGAAGCTAA-3' SEQ ID NO:8.

[0048] (2) The non-transgenic soybean acceptor ZH10, other gene...

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Abstract

The invention discloses a method for quickly identifying the specificity of a transgenic herbicide-resistant soybean ZH10-6 transformant based on a multiplex PCR method. The method comprises the following steps of designing and synthesizing a specific primer by using G2-epsps and GAT exogenous insertion gene sequences of the transgenic herbicide-resistant soybean ZH10-6, and constructing a transformant specificity identification multiplex PCR reaction system through primer concentration optimization combination. Four pairs of transformant specific sequences are designed aiming at a unique expression mode (two repeated G2-epsps genes and one GAT gene are inserted to form two expression cassettes and four boundaries) of an exogenous insertion sequence of the transgenic herbicide-resistant soybean ZH10-6, and a soybean endogenous Lectin gene is introduced to construct a quintuple PCR reaction system. And an economic, efficient, accurate and reliable high-throughput technical means is provided for specific identification of the transgenic transformant based on the exogenous target gene.

Description

technical field [0001] The invention belongs to the technical field of safety evaluation of agricultural transgenic organisms, and in particular relates to the construction and application of a transgenic herbicide-tolerant soybean transformant-specific multiple PCR method. Background technique [0002] With the commercialization of genetically modified crops, the safety of genetically modified crops and their potential threats to human health and the ecological environment have attracted widespread attention from the general public. The "Guidelines for the Safety Evaluation of Genetically Modified Plants" issued by the Ministry of Agriculture of my country provides technical support for the safety evaluation of agricultural GMOs in my country. Among them, molecular characteristics are an important part of the safety assessment of transgenic plants, which refers to the integration, expression and genetic stability of exogenous DNA fragments in the recipient genome in transge...

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Application Information

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IPC IPC(8): C12Q1/6895C12Q1/686C12N15/11
CPCC12Q1/6895C12Q1/686C12Q2600/16C12Q2600/13
Inventor 赵新王一衡李瑞环尉万聪兰青阔王永
Owner 天津市农业科学院
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