ارزیابی توان بیوکنترل و کلنیزاسیون جدایه‌های تریکودرما بهینه‌سازی مولکولی شده در برابر بیماری رایزوکتونیایی لوبیا

نوع مقاله : مقاله پژوهشی

نویسندگان

1 دانشجوی کارشناسی‌ارشد، بخش بیوتکنولوژی گیاهی، پژوهشگاه ملی مهندسی ژنتیک و زیست فناوری، تهران، ایران

2 استاد بخش بیوتکنولوژی گیاهی، پژوهشگاه ملی مهندسی ژنتیک و زیست فناوری، تهران، ایران

3 استادیار بخش بیوتکنولوژی میکروبی، پژوهشگاه بیوتکنولوژی کشاورزی ایران، سازمان تحقیقات، ترویج و آموزش کشاورزی، کرج، ایران

4 کارشناس ارشد بخش بیوتکنولوژی گیاهی، پژوهشگاه ملی مهندسی ژنتیک و زیست فناوری، تهران، ایران

چکیده

کنترل بیولوژیک یکی از راهکارهای مؤثر کنترل بیماری‌های گیاهی و عاملی برای کاهش مصرف سموم شیمیایی است. قارچ تریکودرما با داشتن خاصیت ضدیتی علیه بسیاری از قارچ‌های بیماری‌زای گیاهی از موفق‌ترین عوامل کنترل بیولوژیکی محسوب می‌شود. توان بیوکنترلی این قارچ، با میزان تجزیه آنزیمی دیواره سلولی پاتوژن، ارتباط مستقیم دارد. در پروژه قبلی نویسندگان به کمک مهندسی پروتئین، کیتیناز کایمر 42 ساخته شد و پروتئین نوترکیب حاصل به قارچ T. harzianumانتقال یافت. در تحقیق حاضر توان بیوکنترلی و کلنیزاسیون جدایه‌های مهندسی شده در برابر پاتوژن R. solaniدر شرایط آزمایشگاه و گلخانه بر روی گیاه لوبیا ارزیابی شدند. پس از تأیید مولکولی، حضور و ثبات قطعات مهندسی شده در ریسه قارچ‌های مهندسی شده بعد از گذشت دو سال، بررسی توان بیوکنترلی در کشت متقابل تریکودرما و پاتوژن انجام شد. یافته‌ها نشان داد که جدایه Chit42- ChBD3 با 82/53 درصد بازدارندگی بیش‌تر نسبت به کنترل، بهترین بیوکنترل در شرایط In vitro بوده است. طبق نتایج به‌دست آمده از آزمایش گلخانه‌ای گیاهان تیمار شده با پاتوژن، جدایه‌های مهندسی شده Chit42- ChBD3، Chit42- ChBD7 و Chit42- ChBD15 در تمامی مراحل مختلف اندازه‌گیری (دو برگی، اواسط دوره رویشی، اوایل مرحله زایشی و در مرحله برداشت) علایم بیماری کم‌تراز 40 درصد را نشان دادند. کلنیزاسیون سطحی ریشه و هم‌چنین نفوذ ریسه قارچ به فضای بین سلولی اپیدرم ریشه لوبیا به‌وسیله مطالعات مولکولی با استفاده از آغازگرهای PF1/R3xba1 مورد تائید قرار گرفت.

کلیدواژه‌ها


عنوان مقاله [English]

Evaluation of Bio-control and Colonization Potential of Molecular Improved Trichoderma Strains Against Rhizoctonia Diseases of Bean

نویسندگان [English]

  • Sahar Mahmoodian 1
  • Mohammadreza Zamani 2
  • Mojegan Kowsari 3
  • Mostafa Motallebi 2
  • Esmat Joorabchi 4
1 MSc Student, Department of Plant Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran
2 Professor, Department of Plant Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran
3 Assistant Professor, Department of Microbial Biotechnology and Biosafety, Agricultural Biotechnology Research Institute of Iran (ABRII), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran
4 MSc Graduated, Department of Plant Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran
چکیده [English]

Biological control of plant diseases is one of the effective strategies to reduce harm effects of pesticides on human health and environment. T. harzianum is one of the most successful biological control agents which have antagonistic activity against many of pathogenic fungi. Biocontrol activity of T. harzianum is linked to the amount and type of hydrolase enzymes. In the authors' previous work, chimeric chitinase was constructed by protein engineering and the recombinant protein was transferred to the T. harzianum. In the present study, the biocontrol activity and colonization of improved isolates were evaluated on bean plant and its pathogen (Rhizoctonia solani) in laboratory and greenhouse. The presence and stability of the chimeric protein was confirmed by molecular technique, after that biocontrol assay was done in dual culture method. The results showed that T3 with 82/53 percent inhibition compared to control, was the best biocontrol in invitro. According to the results of greenhouse tests, plants that were treated with T7, T3 and T15 in all stages of measurement (two leaf, mid-term growth, early reproductive stage and harvest stage) showed symptoms less than 40%. Root surface colonization and fungal hyphae penetration was confirmed by molecular studies using primers PF1/R3xba1.

کلیدواژه‌ها [English]

  • Trichoderma
  • Chimeric chitinase
  • Antagonism
  • Protein engineering
  • Pathogen
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