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Primers and detection method for simultaneous detection of cmv, smv and bcmv three viruses in soybean

A CMV-R2 and soybean technology, which is applied in the field of genetic engineering, can solve the problem of not detecting cucumber mosaic virus and common mosaic virus at the same time, and achieve the effects of convenient and quick detection operation, improved detection speed and high sensitivity.

Active Publication Date: 2021-11-16
SICHUAN AGRI UNIV
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Problems solved by technology

The current detection and monitoring methods mainly include conventional biological detection, immunology and molecular biological detection methods, but there are various degrees of defects and deficiencies, and no method has been found that can detect cucumber mosaic virus (CMV) and soybean mosaic virus simultaneously. Method for three viruses of bean mosaic virus (SMV) and bean common mosaic virus (BCMV)

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  • Primers and detection method for simultaneous detection of cmv, smv and bcmv three viruses in soybean
  • Primers and detection method for simultaneous detection of cmv, smv and bcmv three viruses in soybean
  • Primers and detection method for simultaneous detection of cmv, smv and bcmv three viruses in soybean

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Embodiment

[0029] 1. Design primers

[0030] Since the melting temperature between the three pairs of primers may be relatively close, the sequence may appear complementary and other factors, which interfere with the amplification effect. Therefore, after screening, three groups of conserved gene sequences of CMV-870bp, SMV-550bp and BCMV-288bp were selected. Fragment design primers, the specific sequences of which are as follows:

[0031] CMV-F: GGATGCTTCTCCRCGAGDT; (SEQ ID NO. 1)

[0032] CMV-R: GCTGGATGGACAACCCGTTC; (SEQ ID NO. 2)

[0033] SMV-F:AGCTCGCTTCGTCTGGAAAA; (SEQ ID NO.3)

[0034] SMV-R: ATCATCACCCACACGCCATT; (SEQ ID NO.4)

[0035] BCMV-F:GCAACAAAGGAAAAGGYCCTGA; (SEQ ID NO.5)

[0036] BCMV-R: CATACTCGCCCTTCACAGCA. (SEQ ID NO.6)

[0037] 2. Extraction of RNA from soybean leaves

[0038] Take about 0.2 g of soybean leaves, add liquid nitrogen and grind to powder, transfer to Ep tube (placed on ice), add 0.5 mL RNA extraction buffer and 0.5 mL phenol / imitation / isoamyl alc...

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Abstract

The invention discloses a primer for simultaneously detecting three viruses of CMV, SMV and BCMV in soybean and a detection method thereof. The method comprises the following steps: (1) designing primers whose sequences are as shown in SEQ ID NO.1-6; (2) extracting soybean leaf RNA and reverse transcription to prepare cDNA; (3) using the cDNA obtained in step (2) as a template , using the primers described in step (1) for multiplex PCR detection. The primers designed by the invention have strong specificity and high sensitivity, so that the detection speed of soybean virus is greatly improved, especially when the number of samples to be tested is large, it provides a great solution for solving the problems of soybean yield greatly reduced or even crop failure caused by soybean virus. convenience.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a primer for simultaneously detecting three viruses of CMV, SMV and BCMV in soybeans and a detection method thereof. Background technique [0002] Soybean (Glycinemax (L.) Merr) is an important grain and oil crop in my country. According to statistics, 95.526 million tons of soybeans were imported in 2017, and the import dependence rate exceeded 87.7%. It is imminent to increase domestic soybean production. However, in the process of soybean planting, various pests and diseases seriously affect the growth and yield of soybean, among which the losses caused by viruses are very serious. After virus infection, the growth of soybean plants is hindered, the plants are short, the number of pods per plant is reduced, the yield is reduced, and the quality of commodities is reduced. In severe cases, the soybeans have little or no pods, and there are many brown sp...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/701C12Q2600/16C12Q2521/107C12Q2537/143
Inventor 尚静赵兰张磊杨文钰贾琦张敏杨辉孔凡磊常小丽杜俊波余靓雍太文
Owner SICHUAN AGRI UNIV