Fusarium solani Causing Astragalus membranaceus var. mongholicus Root Rot: Establishment of RT-qPCR Detection Method

doi:10.11924/j.issn.1000-6850.casb2023-0496

Abstract

Abstract:

The study aimed at establishing a rapid and accurate real-time fluorescence quantitative PCR (RT-qPCR) method for the detection of Fusarium solani, the pathogenic fungus of Astragalus membranaceus var. mongholicus (AMM) root rot. Based on the elongation factor-1α (EF-1α) sequence of F. solani, a pair of specific primers FP.F/FP.R was designed and verified for its specificity. The RT-qPCR detection method was established and then used to detect the DNA content of F. solani in AMM plants and soil samples for verifying its detection effect. The results showed that the designed specific primer FP.F / FP.R was highly specific, and the sensitivity of the established RT-qPCR method was 0.0192 ng/μL. Repeatability evaluation showed that the method was stable and reliable. With the mothed, the pathogen F. solani was detected in F. solani-inoculated and suspected diseased AMM plants as well as soil samples, the positive detection rate being 100% for all samples in the three groups. Meanwhile, the changes of F. solani content in AMM plants and soil samples were consistent with the severity of root rot. The results indicated that the RT-qPCR method could be used in quantitative detection of F. solani in AMM plants and soil to provide technical support for the early diagnosis of and warning against root rot, and the dynamic monitoring of pathogens.

Key words: Astragalus membranaceus var. mongholicus, root rot, Fusarium solani, real-time fluorescent quantitative PCR, early diagnosis, dynamic monitoring

XU Yingzhi, ZHAO Limei, ZU Weixi, GAO Fen. Fusarium solani Causing Astragalus membranaceus var. mongholicus Root Rot: Establishment of RT-qPCR Detection Method[J]. Chinese Agricultural Science Bulletin, 2024, 40(15): 110-116.

Figures/Tables 6

References 21

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模板/(ng/μL)		Cq值	标准差	变异系数/%
^a不同浓度的腐皮镰刀菌基因组DNA	12	22.88	0.22	0.98
	2.4	25.71	0.34	1.31
	0.48	28.65	0.29	1.00
	0.096	31.49	0.25	0.80
	0.0192	34.51	0.56	1.61
^b不同浓度的腐皮镰刀菌基因组DNA	12	23.50	0.33	1.39
	2.4	26.59	0.32	1.20
	0.48	29.37	0.15	0.53
	0.096	32.01	0.07	0.20
	0.0192	34.57	0.33	0.96

模板/(ng/μL)		Cq值	标准差	变异系数/%
^a不同浓度的腐皮镰刀菌基因组DNA	12	22.88	0.22	0.98
	2.4	25.71	0.34	1.31
	0.48	28.65	0.29	1.00
	0.096	31.49	0.25	0.80
	0.0192	34.51	0.56	1.61
^b不同浓度的腐皮镰刀菌基因组DNA	12	23.50	0.33	1.39
	2.4	26.59	0.32	1.20
	0.48	29.37	0.15	0.53
	0.096	32.01	0.07	0.20
	0.0192	34.57	0.33	0.96

样本类型		样本数	检测部位	阳性样本	腐皮镰刀菌含量/(ng/g)
接种发病样本	0 d	3	病斑	3	10.31±3.58 c
	7 d	3		3	179.02±35.54 b
	14 d	3		3	227.25±20.13 a
	21 d	3		3	223.41±93.68 a
疑似发病样本	Ⅰ	3	芯部	3	46.21±13.15 a
	Ⅱ	3	芯部	3	38.48±8.78 a
	Ⅱ	3	病斑	3	38.87±1.85 a
种植地土壤样本	4年生未发病土	3	根围土	3	32.77±8.63 ab
	4年生发病土	3		3	48.74±3.64 a
	5年生未发病土	3		3	15.53±3.77 b
	5年生发病土	3		3	47.57±18.53 a

样本类型		样本数	检测部位	阳性样本	腐皮镰刀菌含量/(ng/g)
接种发病样本	0 d	3	病斑	3	10.31±3.58 c
	7 d	3		3	179.02±35.54 b
	14 d	3		3	227.25±20.13 a
	21 d	3		3	223.41±93.68 a
疑似发病样本	Ⅰ	3	芯部	3	46.21±13.15 a
	Ⅱ	3	芯部	3	38.48±8.78 a
	Ⅱ	3	病斑	3	38.87±1.85 a
种植地土壤样本	4年生未发病土	3	根围土	3	32.77±8.63 ab
	4年生发病土	3		3	48.74±3.64 a
	5年生未发病土	3		3	15.53±3.77 b
	5年生发病土	3		3	47.57±18.53 a