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Flanking sequence of salt-tolerant transgenic soybean event E8A7027 exogenous insertion fragment and application

A technology of E8A7027 and transgenic soybeans, applied in the field of plant biology, can solve the problem of not being able to effectively realize specific PCR flanking sequences

Inactive Publication Date: 2021-03-26
JILIN ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Based on the flanking sequence of the insertion site of transgenic plants, establishing a specific detection method for transformants and specific detection of transformants and their derivatives is an important technical means to effectively carry out safety supervision of transgenic plants and ensure the healthy development of the transgenic industry. At present, for the salt-tolerant transgenic soybean event E8A7027, there is no flanking sequence that can effectively achieve specific PCR detection

Method used

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  • Flanking sequence of salt-tolerant transgenic soybean event E8A7027 exogenous insertion fragment and application
  • Flanking sequence of salt-tolerant transgenic soybean event E8A7027 exogenous insertion fragment and application
  • Flanking sequence of salt-tolerant transgenic soybean event E8A7027 exogenous insertion fragment and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] In this example, a fragment flanking sequence that can be used for specific PCR detection of the transgenic soybean event E8A7027 was designed based on the analysis of the exogenous insertion site of the transgenic soybean event E8A7027 and the fragment flanking sequence.

[0067] 1.1 Resequencing analysis of the flanking sequence of the exogenous insert in transgenic soybean event E8A7027

[0068] The exogenous insertion site and fragment flanking sequence of the transgenic soybean event E8A7027 were identified by genome resequencing technology. The specific procedure is as follows:

[0069] First, the genomic DNA of the transgenic soybean E8A7027 leaf sample was extracted, and the DNA samples that passed the electrophoresis test were randomly broken into 350bps-500bps DNA fragments by ultrasonic crushing, and the TruSeq DNA LTSample Prep kit was used to build the library. The steps of repairing, adding polyA tails, adding sequencing adapters, purification and PCR amp...

Embodiment 2

[0075] This example provides the specificity of the transgenic soybean event E8A7027 using the 5' flanking sequence (as shown in SEQ ID NO.1) and the 3' flanking sequence (as shown in SEQ ID NO.2) provided in Example 1. The methods of PCR detection include:

[0076] 2.1 Primer design

[0077] The PCR detection primers provided in this example are shown in Table 1.

[0078] Used PCR detection primer in the embodiment 2 of table 1

[0079]

[0080] The primer names 5F, 5R, 3F and 3R respectively correspond to the 5' flanking sequence upstream primer, the 5' flanking sequence downstream primer, the 3' flanking sequence upstream primer and the 3' flanking sequence downstream primer. If the 5' flanking sequence is used as the detection target, the upstream and downstream primers used in step 2.2 are 5F and 5R respectively; if the 3' flanking sequence is used as the detection target, the upstream and downstream primers used in step 2.2 are respectively 3F, 3R.

[0081] 2.2PCR d...

Embodiment 3

[0084] This example provides a method for specific PCR detection of the transgenic soybean event E8A7027 using the 5' flanking sequence (as shown in SEQ ID NO.1) provided in Example 1, and the PCR detection primers used are those described in Example 2 5F and 5R, the detection method is the same as in Example 2.

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Abstract

The invention relates to the technical field of plant biology, in particular to a flanking sequence of a salt-tolerant transgenic soybean event E8A7027 exogenous insertion fragment and an application.The invention discloses a 5'-end flanking sequence and / or 3'-end flanking sequence for exogenous insertion of a salt-tolerant transgenic soybean event E8A7027 for the first time, and designs a primercomposition capable of being used for specific PCR detection according to the 5'-end flanking sequence and / or the 3'-end flanking sequence. The primer composition and the specific PCR detection method can be applied to preparation of a product for detecting a transgenic soybean event E8A7027, so that the safety of the transgenic soybean E8A7027 and a processed product thereof is supervised.

Description

technical field [0001] The invention relates to the field of plant biotechnology, in particular to a flanking sequence of a fragment inserted exogenously into a salt-tolerant transgenic soybean event E8A7027 and its application. Background technique [0002] Soybean is an important grain and oil crop in my country and one of the main raw materials for livestock and poultry feed. In the past 20 years, my country's soybean industry has been severely impacted by imported soybeans. Since 1996, it has changed from a soybean net exporter to a net importer, and the import volume has increased year by year. In 2019, it reached 85.511 million tons, which was 4.72 times the soybean production in my country that year. Soil salinization is one of the important factors affecting soybean yield. Salt stress will lead to a significant decline in key indicators such as soybean plant height, number of main stem nodes, number of pods per plant, and 100-grain weight. The area of ​​saline-alkali...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13
Inventor 李飞武闫伟张玲龙丽坤李葱葱董立明夏蔚邢珍娟刘娜谢彦博何禹璇马月
Owner JILIN ACAD OF AGRI SCI
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