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中国农学通报

中国农学通报 ›› 2025, Vol. 41 ›› Issue (3): 139-144.doi: 10.11924/j.issn.1000-6850.casb2024-0132

• 植物保护·农药 • 上一篇    下一篇

苹果黑星菌LAMP快速检测方法的建立

王华1(), 余海龙2, 任毓忠2()   

  1. 1 伊犁职业技术学院，新疆伊宁 835000
    2 石河子大学农学院/新疆绿洲农业病虫害治理与植保资源利用重点实验室，新疆石河子 832003
  • 收稿日期:2024-02-27 修回日期:2024-05-15 出版日期:2025-01-25 发布日期:2025-01-20
  • 通讯作者:
    任毓忠，男，1971年出生，新疆昌吉人，教授，博士，研究方向：植物病理学。通信地址：832003 新疆石河子市北四路221号，石河子大学，Tel：18999335705，E-mail：。
  • 作者简介:

    王华，女，1973年出生，新疆霍城人，副教授，硕士，研究方向：果树病害防治。通信地址：835000 新疆伊宁市胜利街179号，伊犁职业技术学院，Tel：15886940738，E-mail：。

  • 基金资助:
    新疆维吾尔自治区自然科学基金资助项目“检疫性病害—苹果黑星病病原生物学特性及检测研究”(2020D01A15)

Establishment of Loop-mediated Isothermal Amplification Method for Rapid Detection of Venturia inaequalis

WANG Hua1(), YU Hailong2, REN Yuzhong2()   

  1. 1 Yili Vocational and Technical College, Yining, Xinjiang 835000
    2 College of Agriculture, Shihezi University/Xinjiang Key Laboratory of Pest Control and Plant Protection Resources Utilization in Oasis Agriculture, Shihezi, Xinjiang 832003
  • Received:2024-02-27 Revised:2024-05-15 Published:2025-01-25 Online:2025-01-20

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摘要：

由苹果黑星菌(Venturia inaequalis Cook (Wint.)引起的苹果黑星病是全球苹果重要病害之一，严重影响苹果树的长势、产量及果实品质。为了建立简单、快捷和灵敏的苹果黑星病分子检测方法，以新疆伊犁河谷分离鉴定的苹果黑星菌(V. inaequalis)菌株为试材，基于环介导等温扩增技术(loop-mediated isothermal amplification，LAMP)，以病菌核糖体转录间隔区（ITS区）核酸序列为靶标，设计筛选特异性引物，建立苹果黑星菌的LAMP检测体系，并对其特异性和灵敏性进行验证。结果表明，设计的10组引物中Venin7引物能特异性的从4种苹果病害及1种锈病中鉴定出苹果黑星菌，且检测的稳定性好；该LAMP检测体系在扩增温度55℃~70℃、60~90 min以内均对苹果黑星菌有很好的检测效果；对苹果黑星菌的检测灵敏度为1 pg/μL，比特异性引物PCR检测高出103倍以上。本研究建立的LAMP检测技术，可以快速灵敏地对苹果黑星病进行检测和监测。

关键词: 伊犁河谷, 苹果黑星病, 环介导等温扩增, 病原检测, 病害监测, 防治, 核糖体转录间隔区（ITS区）, LAMP检测体系

Abstract:

Apple scab caused by Venturia inaequalis Cook (Wint.) is one of the important apple diseases worldwide, which seriously affects the growth, yield, and fruit quality of apple trees. In order to establish a simple, fast, and sensitive molecular detection method for apple scab, a strain of apple scab (V. inaequalis) isolated and identified from the Yili River Valley in Xinjiang was used as the test material. Based on loop-mediated isothermal amplification (LAMP) technology, specific primers were designed and screened using the nucleic acid sequence of the ribosome transcription spacer (ITS) region of the pathogen as the target. A LAMP detection system for apple scab was established, and its specificity and sensitivity were verified. The results showed that among the 10 designed primers, the Venin7 primer could specifically identify apple scab from 4 apple diseases and 1 rust disease, and the detection stability was good; the LAMP detection system had good detection performance for apple scab at amplification temperatures of 55℃-70℃ and within 60-90 minutes; the sensitivity of detecting apple scab was 1 pg/μL, which was more than 103 times higher than that of using specific primers for PCR detection. The LAMP detection technology established in this study can quickly and sensitively detect and monitor apple scab.

Key words: Yili River Valley, apple scab, loop-mediated isothermal amplification, pathogen detection, disease monitoring, control, ribosome transcription spacer (ITS) region, LAMP detection system