Unlock instant, AI-driven research and patent intelligence for your innovation.

A kind of plasmid dna standard molecule and application thereof for detecting soybean transgenic components

A technology of genetically modified ingredients and standard molecules, which is applied in the field of plasmid DNA standard molecules to detect genetically modified soybean ingredients, can solve the problems of increasing the workload of detection, pushing up the cost of detection, increasing the workload of DNA extraction, etc., to meet the detection needs Effect

Active Publication Date: 2021-03-05
JILIN ACAD OF AGRI SCI
View PDF7 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the genetically modified screening test, according to the detected target, the genetically modified material containing the corresponding target should be selected as the positive control. Usually, multiple positive controls should be set up for multiple detection targets, which increases the workload of DNA extraction, and in order to examine DNA quality. , it is necessary to set more reactions of internal standard genes, which further pushes up the detection cost, increases the workload of detection, and causes inconvenience to the detection work

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A kind of plasmid dna standard molecule and application thereof for detecting soybean transgenic components
  • A kind of plasmid dna standard molecule and application thereof for detecting soybean transgenic components
  • A kind of plasmid dna standard molecule and application thereof for detecting soybean transgenic components

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Below in conjunction with embodiment the present invention is described in further detail. The following examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. For the experimental methods that do not specify specific conditions in the examples, usually follow the conventional conditions or the conditions suggested by the manufacturer. Embodiment 1, the preparation of the recombinant expression plasmid pUC57-SOY comprising plasmid DNA standard molecule

[0024] 1. Determination of detection targets

[0025] By searching transgenic bioinformatics databases, domestic and foreign patent databases, and detection method databases, the molecular characteristics of 13 transgenic soybean transformants were obtained, including regulatory elements, target genes, marker genes, and transformant-specific sequences. After analyzing the transgenic component information contained in these transformants, the followi...

Embodiment 2

[0038] Embodiment 2, verification of plasmid DNA standard molecule pUC57-SOY

[0039] 1. Activation of recombinant E. coli

[0040] Take out the recombinant Escherichia coli containing the recombinant expression plasmid pUC57-SOY from the -80°C ultra-low temperature refrigerator, put it on ice, do not wait until it melts, directly use the inoculation loop (or pipette tip) to dip the ice in the freezing tube , and then paint the board immediately. Incubate at 37°C for 12-16h. Pick up monoclonal colonies on a sterile operating table, put them into 2-5 mL of LB liquid culture concentrate that has been added with ampicillin antibiotics, and culture them at 37°C and 200 rpm for 8 hours. According to the ratio of 1:500 to 1:1000, dilute the bacterial solution with LB liquid medium with ampicillin antibiotic (25-50μL bacterial solution + 25mL LB liquid medium containing ampicillin), culture at 37°C, 200rpm for 12-16h, To OD600 is about 0.5.

[0041] 2. Sequencing verification

...

Embodiment 3

[0044] Example 3, Application of Recombinant Expression Plasmid pUC57-SOY in Conventional PCR Detection

[0045] 1. Test target

[0046] In order to test the applicability and practicability of the recombinant expression plasmid pUC57-SOY in routine PCR detection, the plasmid DNA was diluted to 500 copy / μL with sterilized deionized water, and 2 μL of the plasmid DNA solution was added to each 25 μL PCR reaction system as a template. PCR amplification. In addition to the detection targets included on pUC57-SOY, some other common targets were selected for amplification, such as internal standard genes of non-soybean crops (maize zSSIIb gene, rice SPS gene, rapeseed HMG gene, cotton Sad-1 gene), Transformant-specific sequences of other transgenic crops (maize transformants MON810, MON863, Bt11, MON89034, MIR604; rice transformants TT51-1, KF-6; cotton transformants MON1445, MON15985; rapeseed transformants GT73, T45), etc., To ensure that no unexpected amplification occurs when...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
PCR efficiencyaaaaaaaaaa
Login to View More

Abstract

The application discloses a plasmid DNA standard molecule for detecting transgene components and a recombinant expression plasmid thereof. The plasmid DNA standard molecule contains the DNA fragment shown in SEQ ID No.2-28. By detecting these targets, most transgenic varieties currently commercialized can be covered. The recombinant expression plasmid of the present invention overcomes the technical problem of lack of positive standard products in transgenic soybean screening detection, gene-specific and transformant-specific qualitative and quantitative PCR detection, and meets the requirement of transgenic soybean component detection. Moreover, it has been verified by experiments that the recombinant expression plasmid has good specificity, no false positive and false negative results are generated, and can be applied commercially.

Description

technical field [0001] The invention relates to the technical field of agricultural transgene detection, in particular to a plasmid DNA standard molecule for detecting soybean transgene components and an application thereof. Background technique [0002] Large-scale commercial planting of genetically modified crops began in 1996. In 2017, the global planting area reached 189.8 million hectares, of which the planting area of ​​genetically modified soybeans was the largest, accounting for 49.6%. There are 41 transgenic soybean transformants approved for marketing in the world. GMO crops have been accompanied by mixed voices since they were first commercialized. On the one hand, genetically modified crops have indeed greatly increased the yield of crops, reduced the use of pesticides, and protected the environment; but on the other hand, the potential uncertainties caused by genetically modified crops to the environment, society and economy have been questioned in many ways. T...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/6895C12Q2600/166C12Q2545/114
Inventor 李飞武闫伟李葱葱龙丽坤董立明刘娜邢珍娟夏蔚谢彦博
Owner JILIN ACAD OF AGRI SCI