基于<em>mtCOI</em>基因的三种短体线虫多重PCR检测

doi:10.3969/j.issn.1674-7968.2020.03.018

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Abstract The Pratylenchus species are important plant-parasitic nematodes and seriously damage the agricultural production worldwide. The establishment of a rapid method for detecting Pratylenchus species is significant to the integrated management of root-lesion nematode diseases. Based on the comparisons of mitochondrial cytochrome oxidaseⅠ(mtCOI) gene sequences from Pratylenchus species, primers were designed, including 3 forward primers Pc-m193-F, Pn-m280-F and Ps-m66-F specific to P. coffeae, P. neglectus and P. scribneri, respectively, and one common reverse primer Pcns-m-R. The PCR products were 225, 137 and 351 bp, respectively, and the PCR sensitivities were 1/1 000, 1/10 000 and 1/100 of individual adult nematode, respectively. The multiplex-PCR system was established successfully after optimizing the concentration ratio of different primers and the annealing temperature. Three Pratylenchus species can be simultaneously and specifically detected using the multiplex-PCR by the combination of primer Pc-m193-F, Pn-m280-F, Ps-m66-F and Pcns-m-R with concentration of 0.4, 0.4, 0.8 and 1.2 μmol/L, respectively, and the annealing temperature with 54 ℃. The detection of 18 Pratylenchus samples using the multiplex-PCR was carried out, and the results were consistent with previous identification. The established multiplex-PCR system could be used to quickly detect the P. coffeae, P. neglectus and P. scribneri either in the samples of single or mixed species infection. This technique could provide the support for diagnosis and integrated management of Triticum aestivum root-lesion nematode diseases in China.

Key words： Multiplex-PCR Mitochondrial cytochrome oxidaseⅠgene (mtCOI) Pratylenchus species Specificity

Received: 30 July 2019

ZTFLH:

S432.4+5

Corresponding Authors: * xuanwang@njau.edu.cn

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	QIN Xin
	WANG Ya-Dong
	LI Hong-Mei
	YU Jia-Rong
	WANG Xuan

Cite this article:

QIN Xin,WANG Ya-Dong,LI Hong-Mei, et al. Multiplex-PCR Detection of Three Pratylenchus Species Based on mtCOI Gene[J]. 农业生物技术学报, 2020, 28(3): 562-570.

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http://journal05.magtech.org.cn/Jwk_ny/EN/10.3969/j.issn.1674-7968.2020.03.018 OR http://journal05.magtech.org.cn/Jwk_ny/EN/Y2020/V28/I3/562

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