Method and special primers for rapidly detecting cowpea thrips sodium ion channel I904S mutation

A sodium ion channel and ion channel technology, applied in the field of rapid detection of the I904S mutation of the cowpea thrips sodium ion channel, can solve the problems of time-consuming, unsuitable for detection, and heavy workload, and achieve the effect of delaying the development of drug resistance

Pending Publication Date: 2021-02-09
HAINAN UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] At present, the traditional method of detecting the mutation site of the sodium ion channel gene of cowpea thrips is to clone the gene, then sequence it and compare it to see if there is a mutation; this method takes a long time due to its heavy workload , it is not suitable for the detection of a large number of samples; the existing technology cannot detect the I904S mutation site of the sodium ion channel gene of the field cowpea thrips in a timely manner

Method used

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  • Method and special primers for rapidly detecting cowpea thrips sodium ion channel I904S mutation
  • Method and special primers for rapidly detecting cowpea thrips sodium ion channel I904S mutation
  • Method and special primers for rapidly detecting cowpea thrips sodium ion channel I904S mutation

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Embodiment 1

[0030] A method for rapid detection of the I904S mutation of the cowpea thrips sodium ion channel and special primers, the primers are a pair: the nucleotide sequence of the upstream primer is shown in SEQ ID NO.2, and the nucleotide sequence of the downstream primer is shown in The nucleotide sequence shown in SEQ ID NO.3; the nucleotide sequence shown in SEQ ID NO.1 is the upstream nucleotide sequence of the thrips cowpea without mutation.

[0031] Utilize above-mentioned primer to identify the method that mutation site I904S of cowpea thrips sodium ion channel gene mutation site is as follows:

[0032] (1) Primer design

[0033] The nucleotide sequences of the primers are shown in Table 1, and allele specific-PCR primers were designed according to the gene sequence of the mutant cowpea thrips sodium ion channel domain II:

[0034] The last base at the 3' end of the dedicated unmutated primer pair is consistent with the base sequence before the mutation (SEQ ID NO.1);

[0...

Embodiment 2

[0048] Sequencing Verification of I904S Mutant Template of Sodium Channel Gene of Vigna thrips

[0049] Specific primers were designed according to the sodium ion channel gene of Vigna grandis thrips (GenBank accession number: MT506055), and the 904-position mutation was amplified by PCR, and the specific target fragment was recovered and sequenced directly.

[0050] PCR amplification primers:

[0051] Upstream primer: SEQ ID NO.2

[0052] Downstream primer: SEQ ID NO.3

[0053] PCR amplification system: pre-denaturation at 94°C for 3 min; denaturation at 94°C for 15 s, annealing at 52°C for 15 s, extension at 72°C for 15 s, and 30 cycles; extension at 72°C for 8 min.

[0054] According to the above, the PCR amplification of the two field populations was completed, and after the agarose gel electrophoresis test analysis, any samples with bands at 270bp were randomly selected for sequencing. The sequencing results are as follows:

[0055] Individual DNA mutation template WT ...

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Abstract

The invention discloses a method and special primers for rapidly detecting cowpea thrips sodium ion channel I904S mutation. The special primers are a pair of detection primers for rapidly identifyingthe cowpea thrips sodium ion channel gene mutation site I904S, and the special primers comprise a first primer with a nucleotide sequence as shown in SEQ ID NO.2 and a second primer with a nucleotidesequence as shown in SEQ ID NO.3. According to the method and the special primers, mutation detection is carried out by using a pair of primers, compared with sequencing, PCR detection is helpful forrapidly and effectively detecting the mutation condition of the mutation site of the field cowpea thrips sodium channel, the real-time state of the drug resistance of the field cowpea thrips to pyrethroids can be rapidly and effectively detected, the technical support is provided for the treatment of drug resistance, and the cowpea thrip prevention and treatment strategy can be conveniently adjusted in time, so that the goal of delaying the development of drug resistance is achieved.

Description

technical field [0001] The invention relates to related technologies and applications thereof in the field of biotechnology, which can monitor the resistance level by detecting the frequency of insect mutations, in particular to a rapid detection method and special primers for the I904S mutation of the Vigna thrips sodium ion channel. Background technique [0002] Vigna thrips is an important pest of leguminous plants. It harms cowpea all the year round in Hainan. Vigna thrips harms plants with adults and larvae. It sucks plant juice through piercing-sucking mouthparts, causing plants to wilt and grow slowly. Hazards and Consequences It can cause suberization of the epidermis, seriously affecting its yield and quality. Cowpea thrips has developed resistance to many insecticides; it has also developed a high level of resistance to pyrethroids; pyrethroids are a class of low-toxicity broad-spectrum Mouthparts play an important role not only in the management of vectors that t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q2531/113C12Q2565/125
Inventor 吴少英潘雪莲李芬袁琳琳陆容材
Owner HAINAN UNIVERSITY
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