Transgenic lettuce (Lactuca sativa L.) harboring chitinase gene expressed resistance against a devastating fungus, Sclerotinia sclerotiorum

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Sharma, Shikha


Research Articles | Published:

Print ISSN : 0970-4078.
Online ISSN : 2229-4473.
Website:www.vegetosindia.org
Pub Email: contact@vegetosindia.org
Doi: 10.1007/s42535-022-00519-8
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Keywords: Lettuce, Transformation, n Chitinase gene, Fungal resistance


Abstract


Lettuce (Lactuca sativa L.) is the world’s most popular leafy salad vegetable cultivated throughout the temperate region of the globe. However, its quality and yield are severely constrained by Sclerotinia sclerotiorum, a destructive fungal pathogen. In this study, rice chitinase (chi) gene was transferred to lettuce cv. Solan Kriti via Agrobacterium-mediated gene transfer technique to obtain resistance against S. sclerotiorum. To enhance the transformation rate, critical determinant variables including explant type, pre-culturing and co-cultivation time and acetosyringone effect were optimized. A combination of 72 h pre-culturing followed by 72 h co-cultivation period resulted in the highest transformation frequency of 8.60% and 3.31% in leaf and petiole explants, respectively. Furthermore, the transformation frequency was enhanced in leaf (23.22%) and petiole (16.44%) explants when the selection medium was fortified with acetosyringone in a concentration of 100 µM, respectively. Transgene (chi) integration was confirmed through PCR and Southern blotting. Expression of the chi gene in transgenic lettuce plants was confirmed by RT-PCR and qRT-PCR analysis. Fungal bioassay of transgenic lettuce proved the effectiveness of chi gene against invasion of Sclerotinia sclerotiorum by showing delay in symptom appearance. Lesions appeared within 12 h after incubation with the fungus in control plants, while leaves of transgenic plant formed lesion after 72 h, but no colonization of the foliar tissue was observed.


Lettuce, Transformation, n                     Chitinase gene, Fungal resistance


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Acknowledgements


Authors are highly thankful to Dr. S. Muthukrishnan, Department of Biochemistry, Kansas state University, Manhattan, USA for providing Agrobacterium strain. The senior author (SS) thankfully acknowledges Department of Science and Technology, New Delhi for providing INSPIRE fellowship during doctoral studies.


Author Information


Sharma Shikha
Department of Biotechnology, Dr. Y.S. Parmar University of Horticulture and Forestry, Solan, India

Gautam Nidhish
RHRSS and KVK, Lahaul-Spiti-II at Tabo, Dr. Y.S. Parmar University of Horticulture and Forestry, Solan, India


Thakur Ajay Kumar
ICAR-Central Potato Research Institute, Shimla, India

thakurak2010@gmail.com
Srivastava D. K.
Department of Biotechnology, Dr. Y.S. Parmar University of Horticulture and Forestry, Solan, India